Fas, FasL, Bcl-2, and Bax in the endometrium of rhesus monkey during the menstrual cycle

To study possible role and regulation of apoptosis occurred in primate endometrium, the expression of apoptosis-related molecules, Fas, FasL, B cell lymphoma/leukaemia-2 (Bcl-2), and Bax were analyzed in relation to occurrence of apoptosis and proliferation in the cycling endometrium of the rhesus monkey using immunohistochemistry and Western blot. The cell apoptosis and proliferation were evaluated by means of in situ 3'-end labeling and Ki67 immunostaining, respectively. The results showed that the expressions of Fas, Fas ligand (FasL), Bcl-2, and Bax were co-localized predominantly in the epithelial cells of the endometrium. Modest Fas staining with no obvious change was detected throughout the menstrual cycle, while the levels of FasL and Bax protein in the epithelial cells increased in the secretory phase when apoptosis was most prevalent. In contrast, epithelial immunostaining for Bcl-2 was maximal during the proliferative phase and decreased in the secretory phase. Bcl-2 immunoreactivity was also detected in some immunocytes. The coordinated expression of Fas, FasL, Bcl-2, and Bax in the cycling endometrium of the rhesus monkey suggests that the cyclic changes in endometrial growth and regression may be regulated by the balance of these factors under the action of ovary steroids.     

Determination of vertebral heart score in three species of Spider monkey (Ateles fusciceps,A. hybridus and A. paniscus)

Background

The vertebral heart score (VHS) is a method of evaluation of cardiac size well documented in domestic mammals and in other primate species, and the aim of this study was to determine the VHS in three species of Spider monkey.

Methods

In this retrospective study, right lateral radiographs of thirty clinically well animals were reviewed and VHS determined. The species included were Ateles fusciceps (n=17), Ateles hybridus (n=8) and Ateles paniscus (n=5).

Results

The VHS was found to vary between species and was 9.73±0.81 for A. fusciceps, 10.53±0.37 for A. hybridus and 10.45±0.27 for A. paniscus.

Conclusions

The observed values appear consistent with values determined for other primate species. There was statistically significant variation noted between species, and so VHS should be considered species‐specific in this genus. The values determined may be of benefit in objectively evaluating cardiac size in the species investigated.     

Role of progesterone on peri-implantation stage endometrium-embryo interaction in the primate

Progesterone secretion during the luteal phase influences oviductal and endometrial functions which are essential for embryo viability and implantation in a number of species including primates. Luteal phase estrogen is not essential for progesterone-dependent endometrial receptivity towards implantation and pregnancy in the rhesus monkey and in the human. However, synchronous development of embryo and endometrium is an essential prerequisite for evolutive implantation. Progesterone helps to maintain synchronous development of preimplantation embryo through its action on maternal uterus. The anti-nidatory action of mifepristone, a potent progesterone receptor modulator (PRM) with pronounced antiprogestagenic activity, is known to be associated with desynchronization of endometrium along with repression of glandular secretory differentiation and vascular maturation. Thus, it is likely that early luteal phase administration of mifepristone affects paracrine action of the secretory stage endometrium on the preimplantation stage embryo, and thereby inhibits embryonic development and viability. We shall examine this hypothesis using the rhesus monkey as a primate model.     

Localization of androgen receptor in the follicle and corpus luteum of the primate ovary during the menstrual cycle

Ovarian androgens may act locally to modulate follicular and luteal function in various species. This study examined the distribution of androgen receptors within the primate ovary throughout the menstrual cycle. Ovaries were collected from rhesus and cynomolgus monkeys during the early, mid-, and late (n = 3-5 per stage) follicular and luteal phases of the cycle. The tissues were processed for indirect immunocytochemical localization of androgen receptors with a specific monoclonal antibody against human androgen receptor (AN1-15). In addition, ovaries (n = 3) were collected from rhesus monkeys for biochemical detection of androgen receptor using 3H-androgen and AN1-15. Specific immunocytochemical staining, as determined by comparing adjacent tissue sections incubated with either AN1-15 or a nonspecific control antibody, was exclusively nuclear. Androgen receptor was detected in the germinal epithelium and ovarian stroma at all stages of the cycle. The thecal and granulosa cells of growing follicles, and of many but not all atretic follicles, contained androgen receptors. Luteinizing granulosa cells of the periovulatory follicle and luteal cells from the early and midluteal phase stained intensely for androgen receptor. Regressing corpora lutea of the late luteal phase also stained for androgen receptor; however, fully regressed corpora lutea in the early follicular phase of the next cycle did not exhibit receptor staining. Luteal cells that were androgen receptor-positive also stained histochemically for the presence of 3 beta-hydroxysteroid dehydrogenase. Sucrose gradient analysis with radiolabeled androgen demonstrated a shift in the androgen receptor peak in monkey ovarian tissue upon addition of AN1-15, confirming the presence of androgen receptor.(ABSTRACT TRUNCATED AT 250 WORDS)     

Molecular cloning of three nonhuman primate follicle stimulating hormone beta-subunit cDNAs

The follicle stimulating hormone (FSH) beta-subunit cDNAs were cloned and sequenced for an old world primate, the rhesus monkey (Macaca mulatta), and two New World primates, the common marmoset (Callithrix jacchus) and pygmy marmoset (Cebuella pygmaea). The cDNA and predicted amino acid sequences of the rhesus monkey FSH beta-subunit were related most closely to the human FSH beta-subunit (> 96% identity). The common and pygmy marmosets have identical FSH beta-subunit cDNAs, whereas the marmoset FSH beta-subunit diverges from the rhesus and human molecules with less than 93% identity. These results have significance for the implementation of assisted reproductive technologies in the nonhuman primate as well as the evolution of genes encoding reproductive hormones.     

Selection of internal reference genes for SYBR green qRT-PCR studies of rhesus monkey (<Emphasis Type="Italic">Macaca mulatta</Emphasis>) tissues

Background  

The rhesus monkey (Macaca mulatta) is a valuable and widely used model animal for biomedical research. However, quantitative analyses of rhesus gene expression profiles under diverse experimental conditions are limited by a shortage of suitable internal controls for the normalization of mRNA levels. In this study, we used a systematic approach for the selection of potential reference genes in the rhesus monkey and compared their suitability to that of the corresponding genes in humans.     

Genome sequence and global sequence variation map with 5.5 million SNPs in Chinese rhesus macaque

Background  

Rhesus macaque (Macaca mulatta) is the most widely used nonhuman primate animal in biomedical research. A global map of genetic variations in rhesus macaque is valuable for both evolutionary and functional studies.     

Immunophenotype and cytokine profiles of rhesus monkey CD56bright and CD56dim decidual natural killer cells

The primate endometrium is characterized in pregnancy by a tissue-specific population of CD56(bright) natural killer (NK) cells. These cells are observed in human, rhesus, and other nonhuman primate decidua. However, other subsets of NK cells are present in the decidua and may play distinct roles in pregnancy. The purpose of this study was to define the surface marker phenotype of rhesus monkey decidual NK (dNK) cell subsets, and to address functional differences by profiling cytokine and chemokine secretion in contrast with decidual T cells and macrophages. Rhesus monkey decidual leukocytes were obtained from early pregnancy tissues, and were characterized by flow cytometry and multiplex assay of secreted factors. We concluded that the major NK cell population in rhesus early pregnancy decidua are CD56(bright) CD16(+)NKp30(-) decidual NK cells, with minor CD56(dim) and CD56(neg) dNK cells. Intracellular cytokine staining demonstrated that CD56(dim) and not CD56(bright) dNK cells are the primary interferon-gamma (IFNG) producers. In addition, the profile of other cytokines, chemokines, and growth factors secreted by these two dNK cell populations was generally similar, but distinct from that of peripheral blood NK cells. Finally, analysis of multiple pregnancies from eight dams revealed that the decidual immune cell profile is characteristic of an individual animal and is consistently maintained across successive pregnancies, suggesting that the uterine immune environment in pregnancy is carefully regulated in the rhesus monkey decidua.     

SERPINE2, an inhibitor of plasminogen activators,is highly expressed in the human endometrium during the secretory phase

Background  

SERPINE2, also known as protease nexin-1, belongs to the serine protease inhibitor (SERPIN) superfamily. It is one of the potent SERPINs that modulates the activity of plasminogen activators (PAs). PAs and their SERPIN inhibitors, such as SERPINB2 and SERPINE1, were expressed in the human endometrium and were implicated in implantation. However, expression data about SERPINE2 in the human endometrium is still unknown. Thus, we conducted an investigation to reveal the spatiotemporal and cellular expression of SERPINE2 in the human uterus during the menstrual cycle.     

Heterochrony and Cross-Species Intersensory Matching by Infant Vervet Monkeys

Background

Understanding the evolutionary origins of a phenotype requires understanding the relationship between ontogenetic and phylogenetic processes. Human infants have been shown to undergo a process of perceptual narrowing during their first year of life, whereby their intersensory ability to match the faces and voices of another species declines as they get older. We investigated the evolutionary origins of this behavioral phenotype by examining whether or not this developmental process occurs in non-human primates as well.

Methodology/Principal Findings

We tested the ability of infant vervet monkeys (Cercopithecus aethiops), ranging in age from 23 to 65 weeks, to match the faces and voices of another non-human primate species (the rhesus monkey, Macaca mulatta). Even though the vervets had no prior exposure to rhesus monkey faces and vocalizations, our findings show that infant vervets can, in fact, recognize the correspondence between rhesus monkey faces and voices (but indicate that they do so by looking at the non-matching face for a greater proportion of overall looking time), and can do so well beyond the age of perceptual narrowing in human infants. Our results further suggest that the pattern of matching by vervet monkeys is influenced by the emotional saliency of the Face+Voice combination. That is, although they looked at the non-matching screen for Face+Voice combinations, they switched to looking at the matching screen when the Voice was replaced with a complex tone of equal duration. Furthermore, an analysis of pupillary responses revealed that their pupils showed greater dilation when looking at the matching natural face/voice combination versus the face/tone combination.

Conclusions/Significance

Because the infant vervets in the current study exhibited cross-species intersensory matching far later in development than do human infants, our findings suggest either that intersensory perceptual narrowing does not occur in Old World monkeys or that it occurs later in development. We argue that these findings reflect the faster rate of neural development in monkeys relative to humans and the resulting differential interaction of this factor with the effects of early experience.     

Chromosome phylogeny of the subfamily Pitheciinae (Platyrrhini,Primates) by classic cytogenetics and chromosome painting

Background  

The New World monkey (Platyrrhini) subfamily Pitheciinae is represented by the genera Pithecia, Chiropotes and Cacajao. In this work we studied the karyotypes of Pithecia irrorata (2n = 48) and Cacajao calvus rubicundus (2n = 45 in males and 2n = 46 in females) by G- and C-banding, NOR staining and chromosome painting using human and Saguinus oedipus whole chromosome probes. The karyotypes of both species were compared with each other and with Chiropotes utahicki (2n = 54) from the literature.     

Establishment and ultrasound characteristics of atherosclerosis in rhesus monkey

Background

Atherosclerosis is one of the main risk factors cause acute cerebral-cardio vascular diseases. It's of great significance to establish an atherosclerosis animal model that can mimic the characteristics and nature course of human patients. Therefore, a rhesus monkey model was induced by high-fat diet to monitor their lipid profile and intima-media thickness (IMT) of artery walls and study atherosclerosis progression.

Methods

Fifty male rhesus monkeys were enrolled in this study. All of these monkeys were aged 7 to 14 years with BMI >30 kg/m². They were fed with high-fat diet containing 10% of fat for the first 48 weeks. Use ultrasound to measure the IMT at bilateral common carotid arteries and their bifurcations and aorta (AO) of the monkeys, and screen out the individuals with thickened IMT for the next phase. In the next 48 weeks, some of these monkeys (n = 4) were fed with standard diet containing 3% fat. Meanwhile the other monkeys (n = 5) were fed with high-fat diet for another 48 weeks. Their serum lipid level was monitored and arterial IMT was also determined periodically.

Results

Serum lipid level of all 50 monkeys elevated after fed with high-fat diet for the first 48 weeks. IMT thickening at right common carotid bifurcation and aorta (AO) was thickened in 9 monkeys. Furthermore, 4 of these 9 monkeys were fed with standard diet and other 5 monkeys were fed with high-fat diet in the following 48 weeks. The serum lipid level of the 4 monkeys recovered and their IMT at RBIF and AO did not progress. However, the lipid level of other 5 monkeys remained high, and their IMT thickening of AO progressed, and plaques and calcification focuses were found at the anterior wall of aorta near the bifurcation of common iliac artery.

Conclusions

After high-fat diet induction for 96 weeks, serum lipid levels of rhesus monkeys elevated significantly, which subsequently caused IMT thickening and plaques formation. When IMT thickening occurred, further vascular injury may be prevented by reducing diet fat content. Our study indicates that vascular injury of high-fat diet induced rhesus monkey is similar to that of human in position and progression.

     

The frequency of CD25+CD4+ and FOXP3+ regulatory T cells in ectopic endometrium and ectopic decidua

Background  

The presence of regulatory T (Treg) cells in human endometrium is crucial for maintaining immunological homeostasis within the uterus. For this study we decided to evaluate the subpopulations of Treg cells in conditions where a disturbance in the immunological equilibrium in ectopic endometrium and decidua has been observed, such as in cases of ovarian endometriosis (involving local immune cell suppression) and ectopic pregnancy (involving an increase in local immune system activity). We then compared these findings to what we observed in the normal eutopic endometrium of women during the secretory phase of the menstrual cycle (with immune cells under individual control).     

Ultrasonographic assessment of the endometrium in rhesus monkeys during the normal menstrual cycle

This study was undertaken to determine whether cyclical changes in the endometrium of the rhesus monkey could be observed by using ultrasound. Three indices of endometrial size were examined: the antero-posterior (or ventro-dorsal), longitudinal, and transverse diameters. Changes in the ultrasonic reflectivity of the endometrium were also assessed. We have attempted to correlate these endometrial parameters with the hormonal status of the animal. Ultrasonography was performed for an average of 12 consecutive days during 19 menstrual cycles. All ultrasonic recordings were normalized to the day of the estradiol (E2) peak (Day 0). We found that the reflectivity of the endometrium was dependent on the stage of the cycle: during the follicular phase, the endometrium appeared less echogenic (darker) compared to the myometrium; in the luteal phase, the endometrium was more echogenic (lighter). During the follicular phase (Days -9 to 0), there was a linear increase in the antero-posterior (p less than 0.001), longitudinal (p less than 0.05), and transverse (p less than 0.001) diameters. In the luteal phase (Days 1-15), no significant changes were observed in these diameters. An estimated endometrial volume (EEV) was obtained by the product of the antero-posterior, longitudinal, and transverse diameters. Each animal observed during the follicular phase (n = 14) exhibited a peak in the EEV, which correlated with the day of the E2 peak (p less than 0.01). From this study, we conclude that the sonographic appearance of the endometrium of the rhesus monkey reflects the cyclical changes that occur during the menstrual cycle.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cloning and Comparison of Factor X from Rhesus Monkey (Macaca mulatta)

The reliability of the rhesus monkey as an important experimental animal depends on its genetic concordance with human. During our assessment of the rhesus monkey as a preclinical model for coagulation-related research, we cloned the full-length cDNA of rhesus monkey factor X (FX) and compared its genetic characteristics and coagulation activity with those of human FX. The full-length cDNA of rhesus monkey FX was 1683 bp in length, corresponding to 487 coding amino acids and sharing 94.71% nucleotide identity and 93.65% amino acid identity with human FX. When FX sequences from different animals were compared with that of human FX, rhesus monkey and baboon FX showed similar degrees of homology to human FX, which were less than that between human and chimpanzee FX sequences but remarkably higher than those of another 2 monkey species, bovine, pig, and rodents. Comparison of functional sites between human and rhesus monkey FX revealed high similarities between their amino acids sequences and 3-dimensional structures. The average coagulation activity of FX from 24 rhesus monkeys was in the normal range of that of healthy humans. The rhesus monkey therefore may be a suitable animal model for research addressing coagulation factor X.Abbreviations: 3D, 3-dimensional; FX, factor X; SMART, simple modular architecture research tool; TFPI, tissue factor pathway inhibitorRhesus monkeys (Macaca mulatta) continue to be widely used research animals in many biologic fields.¹⁵ Because of their close genetic relationship to humans, there is increasing interest in the use of rhesus monkeys for gene therapy, stem cell, infectious disease researches, and reproductive biology.¹² The effective application of rhesus monkey as an important experimental animal is dependent on the genomic and proteomic concordance between rhesus and human.¹⁷ The coagulation system plays an essential role in many physiologic and pathologic processes, including hematologic, cardiovascular, and liver diseases and transplantation. Clearly, the establishment of genetic information and reference activity values of rhesus monkey coagulation factors is requisite to interpretation of the data from preclinical coagulation-related research using rhesus monkeys. Factor X (FX) is a vitamin-K–dependent protein that is 1 of the most critical factors in the coagulation scheme; its activation is the convergence of the extrinsic and intrinsic activation pathways and leads to the final stages of hemostasis.⁶ Human FX has been investigated extensively during the past century, but there have been few reports regarding the characterization of monkey FX.¹⁴The ineffective binding of porcine tissue factor pathway inhibitor (TFPI) to human FX was proposed to be an important contributor to coagulopathy in pig-to-human xenotransplantation.¹¹ Many in vitro studies¹¹ have investigated the interaction between porcine endothelial cells and human coagulation factors, whereas in vivo studies⁸ have addressed the interaction between porcine endothelial cells and monkey coagulation factors. The concordance of the results from these in vitro and in vivo studies therefore reflects high homology, if not identity, between human FX and monkey FX. During our assessment of whether rhesus monkey is a reliable model for studying coagulation disorders in xenotransplantation, we cloned the full-length cDNA of rhesus monkey FX and compared its nucleotide and amino-acid sequences and coagulation activities with those of human FX.     

IL-11 and IL-11Rα immunolocalisation at primate implantation sites supports a role for IL-11 in placentation and fetal development

Embryo implantation, endometrial stromal cell decidualization and formation of a functional placenta are critical processes in the establishment and maintenance of pregnancy. Interleukin (IL)-11 signalling is essential for adequate decidualization in the mouse uterus and IL-11 promotes decidualization in the human. IL-11 action is mediated via binding to the specific IL-11 receptor α (IL-11Rα). The present study examined immunoreactive IL-11 and IL-11Rα in cycling rhesus monkey endometrium, at implantation sites in cynomolgus and rhesus monkeys and in human first trimester decidua and defined distinct spatial and temporal patterns. In cycling rhesus monkey endometrium, IL-11 and IL-11Rα increased in both basalis and functionalis regions during the secretory compared with the proliferative phase, with changing cellular locations in luminal and glandular epithelium and stroma. The patterns were similar overall to those previously described in human endometrium. Differences were seen in immunostaining during implantation in cynomologus and rhesus monkey. In the cynomolgus, very little staining for IL-11 or IL-11Rα was seen in syncytio- and cyto-trophoblast cells in the villi between days 12 and 150 of pregnancy although there was moderate staining in cytotrophoblast in the shell between days 12 and 17 and in subpopulations of cytotrophoblast cells invading the arteries at day 17. By contrast in the rhesus monkey between days 24 and 35 of pregnancy and in human first trimester placenta, cyto- and syncytio-trophoblast in the villi but not cytotrophoblast in the shell were positively stained. The most intense staining for both IL-11 and IL-11Rα was present within the decidua in the maternal component of implantation sites in all three primates but moderate staining was also present in maternal vascular smooth muscle and glands perivascular cells and epithelial plaques. These results are consistent with a role for IL-11 both during decidualization and placentation in primates.     

Autoradiographic localization of aromatic hydrocarbon receptor (AHR) in rhesus monkey ovary

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is the most toxic congener of a large class of manmade pollutants that persist in the environment. TCDD exerts its toxic effects, in part, by binding to its receptor known as the aromatic hydrocarbon receptor (AHR). TCDD is estrogen modulatory and in some systems its receptor associates directly with estrogen receptors via co-activator molecules. TCDD inhibits steroid synthesis in human ovarian granulosa cells and AHR is found in these cells. We have previously shown that AHR is found in whole rhesus monkey ovary, but have yet to establish its location. In the present study, we set out to show that radiolabeled TCDD binds to monkey ovarian follicles and that this binding is receptor mediated. Ovaries from Macaca mulatta were sectioned on a cryostat at 10 micro m; and sections were incubated with either control vehicle, (3)H-TCDD, or (3)H-TCDD plus alpha-naphthoflavone (ANF), a known receptor-blocking agent. Here, we show for the first time specific binding of TCDD to the granulosa cells of antral follicles and other regions of the rhesus monkey ovary. Our data indicate a 60-fold increase in binding with (3)H-TCDD over that of control, and that this binding is reduced to the levels seen in controls with the addition of the competitive antagonist ANF. These findings support the hypothesis that TCDD directly affects primate ovarian function via the AHR.