Embryotoxicity of sex steroidal hormone combinations in nonhuman primates: I. Norethisterone acetate + ethinylestradiol and progesterone + estradiol benzoate (Macaca mulatta, Macaca fascicularis, and Papio cynocephalus)

Two sex steroid hormone combinations which have been used clinically as tests for detection of early pregnancy were examined for embryotoxic effects in macaques and baboons. Norethisterone acetate and ethinyl estradiol (NEA + EE) were orally administered to rhesus and cynomolgus monkeys and baboons at dosages ranging from one to 1,000 times the human dose equivalent (HDE) during days 20-50 of pregnancy. Progesterone and estradiol benzoate (P + EB) were delivered by two to six intramuscular injections to rhesus and cynomolgus monkeys between gestational days 20 and 35 at 0.1-25 X HDE. Fetuses were examined following cesarean section at 100 +/- 2 days (NEA + EE) or at term (P + EB). The results showed increased embryolethality over controls at 100-1,000 X HDE (NEA + EE) and at 10 and 25 X HDE (P + EB). Besides growth retardation, isolated cases of minor nongenital malformations were observed only in cynomolgus monkeys following treatment with both hormone combinations mainly at embryolethal dose levels and were considered spontaneous in nature. Virilization of female cynomolgus fetuses following NEA + EE treatment was manifested as two cases of clitoral enlargement in the 300 X HDE group and two cases of increased anogenital distance with reduced vaginal opening in the 1,000 X HDE group. The highest dose of NEA + EE was also maternally toxic, as two maternal deaths occurred at the end of the treatment period. One dead female cynomolgus fetus exposed to P + EB (10 X HDE) also exhibited masculinized external genitalia.(ABSTRACT TRUNCATED AT 250 WORDS)     

Evaluation of bendectin embryotoxicity in nonhuman primates: I. Ventricular septal defects in prenatal macaques and baboon

Cynomolgus monkeys, rhesus monkeys and baboons were administered 10 to 40 times the human dose equivalent of Bendectin throughout the major period of organogenesis (22(+/-3)-50 days of gestation). In animals examined prenatally (100 +/- 2 days gestation) the total incidence of ventricular septal defects (VSD) was 40% in cynomolgus monkeys, 18% in rhesus monkeys, and 23% in baboons. The majority of VSD involved the muscular portion of the septum. No dose response was evident and there were no other cardiac or extracardiac defects found except for one baboon fetus with multiple defects. No defects were observed in cynomolgus monkeys administered Bendectin for 4-day periods between 22 and 41 days of gestation. There was no association of Bendectin treatment with any noncardiac defect. In cynomolgus and rhesus monkeys examined at term there was one mitral valve defect and no incidence of VSD. The increased incidence of VSD observed prenatally in all three species and the absence of defects in macaques at term suggests a delay in closure of the ventricular septum in treated animals. The Bendectin-treated monkey may be a suitable model for the study of the pathogenesis of VSD and the mechanism of spontaneous closure of the defect.     

Movement of cynomolgus and rhesus monkey spermatozoa collected from the lower female reproductive tract

Postcoital (pc) cervical mucus was collected in 73 menstrual cycles of cynomolgus monkeys and in 43 cycles of rhesus monkeys at 2,6,10,30 hr pc. Videomicrography was used to analyze sperm numbers and movement in the mucus. Both cynomolgus and rhesus monkeys had comparable populations of motile sperm in the mucus at 2 hr pc. However, by 6 hr pc, cervical mucus from cynomolgus monkeys contained twice as many total sperm and motile sperm as mucus from rhesus monkeys (P <.05). Mean swimming speeds of the free-swimming cervical sperm were similar for the two species at this time. No motile sperm were recovered in mucus from rhesus monkeys at 30 hr pc. In cynomolgus monkeys, however, 14 of the 26 animals examined at 30 hr pc had motile sperm in their mucus. These sperm exhibited lower percent molility, percent free-swimming sperm, and swimming speed than those sperm observed at 6 hr pc. Uterine sperm were collected by transcervical or transuterine aspiration from cynomolgus monkeys. In the transcervical technique, sperm were successfully obtained in four of nine animals examined at 6 hr and in four of five animals at 30 hr pc. The percentage of motile sperm in the uterine fluid was high, 82% ± 4%, and the swimming speeds (86 ± 2μm/sec) were higher than those observed in cervical mucus. Approximately 5–10% of the uterine sperm exhibited swimming motions similar to the hyperactivated motility seen in most mammals. These findings indicate that the sperm cervical mucus interaction in vivo in cynomolgus monkeys has more similarities to the human situation than does the interaction in rhesus monkeys.     

Effect of gestational sex steroid exposure on limb development and endochondral ossification in the pregnant C57Bl/6J mouse: I. Medroxyprogesterone acetate

Although data supporting the teratogenic potential of intrauterine progestin exposure is lacking, concern persists among some individuals within the scientific community that these drugs have the potential for nongenital teratogenesis, especially with regard to limb reduction defects. Our laboratory has been interested in the ontogeny of steroid receptors in the developing embryo and in the role of steroid-receptor interactions in limb development, particularly the process of endochondral ossification. Since limb reduction defects can be produced from abnormal processes that are operative during organogenesis or during midgestation (vascular disruption) we have designed an animal study whereby embryos were exposed to sex steroids throughout organogenesis and fetal development. The present study assesses the effects of medroxyprogesterone acetate (MPA) on intrauterine endochrondral bone development specifically, as well as overall embryo-fetal development. Primagravid C57Bl/6J mice were treated via subdermal pellets which deliver MPA at dosages of 5.0, 50.0, and 500.0 mg/kg/day on gestational days 7 through 19. These doses were 25-, 250-, and 2,500-fold higher on a mg/kg basis than the human dose equivalent (HDE). No increases in nongenital malformations were noted at any evaluated MPA dosage level. At 25 X the HDE, MPA did not influence endochondral bone development as evidenced by a lack of significant effects on assessed bone growth parameters. In the 250- and 2,500-fold HDE dosage groups, MPA was shown to exert an embryotoxic effect inducing 48 and 100% resorptions respectively. Mean embryo weights/litter were significantly reduced by MPA exposure at 250 X the HDE. Intrauterine exposure to 250 X the MPA HDE induced reductions in humeral and femoral diaphyseal length in proportion to a reduction in overall growth. The data demonstrate that MPA, administered at dosages of up to several orders of magnitude in excess of the HDE and which permitted embryo survival, did not induce increases in the frequency of nongenital teratogenesis at any dose or gestational stage. Importantly, limb reduction defects were not noted even in instances where the dosage of MPA induced an inhibition of endochondral bone growth.     

Effects of testosterone on the behavior of male cynomolgus monkeys (Macaca fascicularis)

To determine the threshold doses of testosterone propionate (TP) that cause clear-cut behavioral changes in the sexual behavior of castrated male cynomolgus monkeys, observations were made on three males during successive 5-week treatment periods while they received daily subcutaneous doses of 100 μg TP increasing in octaves to 25.6 mg TP. Males were tested with each of the same two ovariectomized, estrogen-treated females (6 pairs, 330 1-hr behavior tests). To mimic the diurnal plasma testosterone rhythm, TP injections were given at 1600 hr and blood samples were obtained at 0800 hr (141 samples). Male ejaculatory activity increased at the threshold dose of 200 μg TP per day giving plasma testosterone levels of 830 ng/100 ml, which is in the physiological range of 600–1600 ng/100 ml for intact males. This threshold dose was eight times higher than in rhesus monkeys on a dose per kilogram body weight basis. There was a further marked increase in ejaculatory performance at higher doses (6.4 to 25.6 mg) giving supraphysiological plasma levels of 4000–9000 ng/100 ml. There were individual differences in the behavioral changes occurring with TP treatment, and the female partner modulated the effects. These findings were generally similar to those obtained with male rhesus monkeys, but certain species differences were noted.     

Aerosol exposure to Zaire ebolavirus in three nonhuman primate species: differences in disease course and clinical pathology

There is little known concerning the disease caused by Zaire ebolavirus (ZEBOV) when inhaled, the likely route of exposure in a biological attack. Cynomolgus macaques, rhesus macaques, and African green monkeys were exposed to aerosolized ZEBOV to determine which species might be the most relevant model of the human disease. A petechial rash was noted on cynomolgus and rhesus macaques after fever onset but not on African green monkeys. Fever duration was shortest in rhesus macaques (62.7 ± 16.3 h) and longest in cynomolgus macaques (82.7 ± 22.3 h) and African green monkeys (88.4 ± 16.7 h). Virus was first detectable in the blood 3 days after challenge; the level of viremia was comparable among all three species. Hematological changes were noted in all three species, including decreases in lymphocyte and platelet counts. Increased blood coagulation times were most pronounced in African green monkeys. Clinical signs and time to death in all three species were comparable to what has been reported previously for each species after parenteral inoculation with ZEBOV. These data will be useful in selection of an animal model for efficacy studies.     

Biologic data of Macaca mulatta, Macaca fascicularis, and Saimiri sciureus used for research at the Fiocruz primate center

Physiological parameters of laboratory animals used for biomedical research is crucial for following several experimental procedures. With the intent to establish baseline biologic parameters for non-human primates held in closed colonies, hematological and morphometric data of captive monkeys were determined. Data of clinically healthy rhesus macaques (Macaca mulatta), cynomolgus monkeys (Macaca fascicularis), and squirrel monkeys (Saimiri sciureus) were collected over a period of five years. Animals were separated according to sex and divided into five age groups. Hematological data were compared with those in the literature by Student's t test. Discrepancies with significance levels of 0.1, 1 or 5% were found in the hematological studies. Growth curves showed that the sexual dimorphism of rhesus monkeys appeared at an age of four years. In earlier ages, the differences between sexes could not be distinguished (p < 0.05). Sexual dimorphism in both squirrel monkeys and cynomolgus monkeys occurred at an age of about 32 months. Data presented in this paper could be useful for comparative studies using primates under similar conditions.     

Evaluation of cynomolgus (Macaca fascicularis) and rhesus (Macaca mulatta) monkeys as experimental models of acute Q fever after aerosol exposure to phase-I Coxiella burnetii

BACKGROUND AND PURPOSE: Q fever is a disease of humans. Vaccines to prevent this disease have demonstrated efficacy in rodents and must also be evaluated for efficacy in a nonhuman primate model. Preliminary to vaccine efficacy experiments, cynomolgus and rhesus monkeys were evaluated as suitable experimental models of acute Q fever. METHODS: Both species of monkeys were challenged with aerosolized 10(5) virulent phase-I Coxiella burnetii Henzerling strain, and clinical and serologic responses were determined. RESULTS: Radiographic changes were observed in seven of eight monkeys of both species; however, changes in cynomolgus monkeys tended to be more significant. Between 7 and 10 days after challenge, all rhesus monkeys and 88% of cynomolgus monkeys were bacteremic. Sequential increases in antibody responses to C. burnetii phase-I and phase-II whole cells and phase-I lipopolysaccharide were observed in both species. Although the maximal rectal temperature increase was similar in both species, duration of fever was slightly longer in rhesus monkeys. Clinical features were similar to those described in human acute Q fever patients. CONCLUSIONS: On the basis of the more pronounced radiographic changes in cynomolgus monkeys, we favor use of this species for future studies of vaccine efficacy.     

Human fetuses do not register chromosome damage inflicted by radiation exposure in lymphoid precursor cells except for a small but significant effect at low doses

Human fetuses are thought to be highly sensitive to radiation exposure because diagnostic low-dose X rays have been suggested to increase the risk of childhood leukemia. However, animal studies generally have not demonstrated a high radiosensitivity of fetuses, and the underlying causes for the discrepancy remain unidentified. We examined atomic bomb survivors exposed in utero for translocation frequencies in blood lymphocytes at 40 years of age. Contrary to our expectation of a greater radiosensitivity in fetuses than in adults, the frequency did not increase with dose except for a small increase (less than 1%) at doses below 0.1 Sv, which was statistically significant. We interpret the results as indicating that fetal lymphoid precursor cells comprise two subpopulations. One is small in number, sensitive to the induction of both translocations and cell killing, but rapidly diminishing above 50 mSv. The other is the major fraction but is insensitive to registering damage expressed as chromosome aberrations. Our results provide a biological basis for resolving the long-standing controversy that a substantial risk of childhood leukemia is implicated in human fetuses exposed to low-dose X rays whereas animal studies involving mainly high-dose exposures generally do not confirm it.     

Embryotoxicity studies of norfloxacin in cynomolgus monkeys: I. Teratology studies and norfloxacin plasma concentration in pregnant and nonpregnant monkeys

Norfloxacin, a new orally active antibiotic, was investigated in cynomolgus monkeys for potential developmental toxicity. Fifty-seven monkeys were administered a control vehicle or norfloxacin by nasogastric gavage during the major period of organogenesis on gestational days (GD) 21 through 50 at doses of 0, 50, 100, 150, or 200/300 mg/kg/day. There was no evidence of teratogenicity at any dose level. Maternotoxicity and a significant increase in embryolethality occurred following doses of 200/300 mg/kg/day. The maternotoxicity was not expected based on range-finding studies in nonpregnant female monkeys, which showed no signs of toxicity in doses up to 500 mg/kg/day. Additional studies were conducted to determine if norfloxacin caused similar toxicity later in gestation. Forty-six pregnant monkeys were dosed with a control vehicle or 200 mg/kg/day norfloxacin for one of three 10-day periods on GD 36-45, 71-80, or 111-120. There were no maternotoxic, embryotoxic, or fetotoxic effects observed. Plasma concentrations of norfloxacin in five cynomolgus monkeys following 50 and 200 mg/kg oral doses were not dose-proportionate. However, at a given dose, administered in cross-over fashion, plasma concentrations of norfloxacin were higher in nonpregnant females (approximately 20-40%) than during pregnancy when the same subject was compared. At the no-observed-effect dose for maternal and embryotoxicity (50 mg/kg), peak plasma concentrations of norfloxacin in pregnant cynomolgus monkeys are approximately threefold higher than those observed in human volunteers receiving norfloxacin at the maximum recommended therapeutic dose of 400 mg (5.7 mg/kg based on 70 kg body weight) twice per day.     

人工饲养恒河猴、食蟹猴的繁殖性能初报

目的探索北京地区人工饲养恒河猴与食蟹猴的繁殖性能,为温带地区猕猴的人工饲养和繁殖方式提供借鉴。方法对军事医学科学院实验动物中心饲养的317只恒河猴繁殖群（30只雄猴,287只雌猴）和78只食蟹猴繁殖群（8只雄猴,70只雌猴）近两年的繁殖性状进行观察和统计分析。结果恒河猴母猴妊娠率、繁殖率和成活率分别为60.73%、54.45%和96.89%。食蟹猴母猴妊娠率、繁殖率和成活率分别为79.86%、56.12%和75.00%。结论食蟹猴和恒河猴可以成功的在温带地区饲养和繁殖,但人工饲养食蟹猴的妊娠率与产仔率较恒河猴高,而仔猴成活率则低于恒河猴。     

Distribution and biological effects of inhaled 239Pu(NO3)4 in cynomolgus monkeys.

Twenty male cynomolgus monkeys were exposed by inhalation either to an aerosol of 239Pu(NO3)4 to produce projected initial lung burdens of either 40, 10, or 4 kBq or to a carrier aerosol as a control. Animals died or were sacrificed at 0.01, 1, 3, 6, 12, 24, 40, and 99 months after inhalation, and the distribution and biological effects of the 239Pu were determined. The 239Pu cleared efficiently from the lungs so that less than 0.05 kBq remained at 99 months after exposure to 40 kBq. Total skeletal 239Pu activity was nearly constant after the first year, but the fraction of the body burden in skeleton at sacrifice increased with time up to 99 months because of clearance from other organs. Plutonium in the liver increased to a peak at 1 year and then decreased to about 10% of the peak value at 99 months. Plutonium in the testes was localized in the interstitial tissue with only 0.01 to 0.002% of the projected lung burden remaining in testes at 99 months after inhalation. Three animals exposed to 40 kBq of 239Pu died of radiation-related pulmonary pneumonitis and fibrosis. A primary papillary adenocarcinoma of the lung was identified in one animal exposed to 40 kBq initial lung burden and sacrificed 99 months after inhalation. The frequency of chromosome aberrations in blood lymphocytes was significantly elevated only in monkeys with projected deposits of 40 kBq of 239Pu. There was no change in aberration frequency in other exposure groups as a function of inhaled activity, time after exposure, or calculated total dose to the lungs. Only in monkeys that had marked radiation-induced pathological changes in the lung did the frequency of chromosome-type aberrations increase significantly, to a value about twice the control level. In cynomolgus monkeys, chromosome aberration frequency in blood lymphocytes is not a good indicator of radiation dose or damage from inhaled soluble plutonium.     

Comparative studies with six extenders for sperm cryopreservation in the cynomolgus monkey (Macaca fascicularis) and rhesus monkey (Macaca mulatta)

Ejaculated spermatozoa from cynomolgus monkeys and rhesus monkeys were frozen in straws with six different extenders (TTE, DM, mDM, LG-DM, G-DM, and TCG) containing glycerol. Sperm motility and head membrane and acrosomal integrity were evaluated after freezing and thawing, and the cryoprotective effects were compared among the extenders and the two species studied. The results showed that sperm motility and motility recovery with the six extenders were comparable for the cynomolgus and rhesus monkeys. There was no significant difference in sperm motility and head membrane integrity among the six extenders in either the cynomolgus or rhesus monkeys (P>0.05). However, a slightly but statistically lower percentage of acrosomal integrity was found with TCG in both species compared to the other extenders (P<0.05). These findings demonstrate that TTE, DM, mDM, LG-DM, G-DM, and TCG are equally suitable extenders for the cryopreservation of spermatozoa from cynomolgus and rhesus monkeys.     

Mutational extinction induced by sequential X irradiation and actinomycin D treatments in Chinese hamster ovary cells

The interactions of sequential X irradiation and actinomycin D (AMD) treatments for mutagenesis to 6-thioguanine resistance were investigated in CHO cells. Cells were exposed to single doses of X rays followed immediately by 1-h treatments with 0.1 or 1 microgram/ml AMD. X Rays alone induced mutagenesis which increased monotonically with dose to at least 8 Gy. AMD-treated control cultures showed slight to moderate cytotoxicity and little induced mutation. X Rays followed by AMD treatment produced bell-shaped mutagenesis dose-response curves with maximal mutation at approximately 5 or 4 Gy for 0.1 or 1.0 microgram/ml AMD, respectively. Induced mutation frequencies then fell to a negligible level at fractional survival levels below 0.10 for either combination treatment. Application of a stochastic Poisson distribution model to these data led to the prediction that two possible components govern induced mutation frequencies. First, X ray +AMD induced mutations may be depleted progressively with dose from the surviving populations by selective lethality, which we term mutational extinction. Second, X ray +AMD treatments were calculated to induce potentially much greater than additive mutagenesis. However, due to the overriding mutational extinction effect, most of these mutations are not recovered as viable colonies. These studies suggest that AMD binding to DNA immediately following irradiation may cause considerably enhanced mutagenic and often lethal DNA damage, and that mutational extinction may occur because these types of damage are statistically correlated in a sensitive subpopulation of exponentially growing CHO cells.     

Preclinical pharmacokinetics of MHAA4549A,a human monoclonal antibody to influenza A virus,and the prediction of its efficacious clinical dose for the treatment of patients hospitalized with influenza A

MHAA4549A is a human immunoglobulin G1 (IgG1) monoclonal antibody that binds to a highly conserved epitope on the stalk of influenza A hemagglutinin and blocks the hemagglutinin-mediated membrane fusion in the endosome, neutralizing all known human influenza A strains. Pharmacokinetics (PK) of MHAA4549A and its related antibodies were determined in DBA/2J and Balb-c mice at 5 mg/kg and in cynomolgus monkeys at 5 and 100 mg/kg as a single intravenous dose. Serum samples were analyzed for antibody concentrations using an ELISA and the PK was evaluated using WinNonlin software. Human PK profiles were projected based on the PK in monkeys using species-invariant time method. The human efficacious dose projection was based on in vivo nonclinical pharmacological active doses, exposure in mouse infection models and expected human PK. The PK profiles of MHAA4549A and its related antibody showed a linear bi-exponential disposition in mice and cynomolgus monkeys. In mice, clearance and half-life ranged from 5.77 to 9.98 mL/day/kg and 10.2 to 5.76 days, respectively. In cynomolgus monkeys, clearance and half-life ranged from 4.33 to 4.34 mL/day/kg and 11.3 to 11.9 days, respectively. The predicted clearance in humans was ～2.60 mL/day/kg. A single intravenous dose ranging from 15 to 45 mg/kg was predicted to achieve efficacious exposure in humans. In conclusion, the PK of MHAA4549A was as expected for a human IgG1 monoclonal antibody that lacks known endogenous host targets. The predicted clearance and projected efficacious doses in humans for MHAA4549A have been verified in a Phase 1 study and Phase 2a study, respectively.     

Viremia and antibody response in green monkeys (Chlorocebus aethiops sabaeus) infected with dengue virus type 2: A potential model for vaccine testing

The increasingly limited availability and high cost of the hitherto most commonly used monkey species in dengue vaccine research has augmented the importance of identifying alternative suitable models for these studies. In this study we examined the capacity of green monkeys ( Chlorocebus aethiops sabaeus ) to develop dengue viremia, and thus provide a potential model for dengue vaccine testing. Monkeys were inoculated with two different doses of dengue virus type 2. All animals in both groups became viremic after inoculation of the virus. In the lower dose group, mean viremia duration of 5.66 days was detected, whereas in the group that received the 10⁶ PFU dose, viremia had a mean duration of only 1.66 days. Antibody titers were similar to those obtained in previous experiments with rhesus and cynomolgus macaques. We conclude that green monkeys develop viremia and antibody responses and therefore provide a potential model for the preclinical evaluation of novel candidates for dengue vaccines.     

An attempt to predict anergy in tuberculosis suspect cynomolgus monkeys.

Acid-fast microorganisms were identified from the tuberculous lesions of a male cynomolgus monkey (Macaca fascicularis). Twenty-two other cynomolgus monkeys housed in the same room were presumed exposed to tuberculosis (Mycobacterium spp.). In addition to standard intradermal (ID) tuberculin testing, clinicians attempted to evaluate the immune status of these monkeys in order to identify animals exhibiting false negative (anergy) ID tuberculin tests. Twenty-one of the potentially exposed monkeys were immunized with tetanus toxoid (TT). Tetanus antitoxin (TAT) titers were measured before and after immunization. The delayed cutaneous hypersensitivity (DCH) reaction to TT was evaluated using a commercially available human test panel. Some animals did not exhibit a DCH reaction to TT. At necropsy 1 of the 21 animals exhibited tuberculous lesions, and acid-fast microorganisms were identified on direct smears of lymphatic tissue of a second animal. Although reported to be of value in assessing the cellular immune status of rhesus monkeys (Macaca mulatta), the delayed cutaneous hypersensitivity response to tetanus toxoid was not helpful during this outbreak in identifying cynomolgus monkeys infected with M. tuberculosis, or in interpreting suspect ID tuberculin tests.     

Nickel and Ultraviolet-B Stresses Induce Differential Growth and Photosynthetic Responses in Pisum sativum L. Seedlings

Enhanced level of UV-B radiation and heavy metals in irrigated soils due to anthropogenic activities are deteriorating the environmental conditions necessary for growth and development of plants. The present study was undertaken to study the individual and interactive effects of heavy metal nickel (NiCl(2)·6H(2)O; 0.01, 0.1, 1.0?mM) and UV-B exposure (0.4 W m(-2); 45?min corresponds to 1.08 KJ m(-2)) on growth performance and photosynthetic activity of pea (Pisum sativum L.) seedlings. Ni treatment at high doses (0.1 and 1.0?mM Ni) and UV-B alone reduced chlorophyll content and photosynthetic activity (oxygen yield, carbon fixation, photorespiration, and PSI, PSII, and whole chain electron transport activities), and declining trends continued with combined doses. In contrast to this, Ni at 0.01?mM appeared to be stimulatory for photosynthetic pigments and photosynthetic activity, thereby enhanced biomass was observed at this concentration. However, combined dose (UV-B + 0.01?mM Ni) caused inhibitory effects. Carotenoids showed different responses to each stress. Nickel at high doses strongly inhibited PSII activity and the inhibition was further intensified when chloroplasts were simultaneously exposed to UV-B radiation. PSI activity appeared to be more resistant to each stress. High doses of Ni (0.1and 1.0?mM) and UV-B alone interrupted electron flow at the oxygen evolving complex. Similar damaging effects were caused by 0.01 and 0.1?mM Ni together with UV-B, but the damage extended to PSII reaction center in case of 1.0?mM Ni in combination with UV-B. In conclusion, the results demonstrate that low dose of Ni stimulated the growth performance of pea seedlings in contrast to its inhibitory role at high doses. However, UV-B alone and together with low as well as high doses of Ni proved to be toxic for P. sativum L.     

Development of a microsatellite marker set applicable to genome-wide screening of cynomolgus monkeys (<Emphasis Type="Italic">Macaca fascicularis</Emphasis>)

To develop a microsatellite marker set applicable to genome-wide screening of cynomolgus monkeys (Macaca fascicularis), 148 microsatellite markers were selected from the human genome database. The polymorphisms and inheritance of PCR products were determined by screening twenty unrelated monkeys and by analysis of three families, respectively. As a result, 106 primers (72%) gave PCR products of the size expected for humans and rhesus monkeys. Among these products, polymorphism and single-gene inheritance in cynomolgus monkeys was observed for 66 markers (62%). The average number of alleles at the 66 polymorphic loci was 5.86 (range 2–10), and average heterozygosity was 0.63 (range 0.10–0.88). This is the first report of microsatellite markers for cynomolgus monkeys. Chromosomal mapping of these markers is now in progress.     

Synergism between psychosocial and metabolic stressors: impact on reproductive function in cynomolgus monkeys

The role of energy imbalance versus psychosocial stress in the pathogenesis of female reproductive dysfunction characterized by anovulation and amenorrhea remains controversial. In women, functional hypothalamic amenorrhea can develop in the absence of significant weight loss, excessive exercise, or profound psychosocial disruption. We posited, therefore, that commonplace, seemingly minor stressors that alone would have minimal impact upon reproductive function might interact synergistically such that combinations of stressors would cause a greater impairment of the reproductive axis than any single stressor alone. We then developed a monkey model to test this hypothesis. Adult female cynomolgus monkeys with normal menstrual cycles were randomized into three experimental groups and studied over four menstrual cycles. The groups were: low-level psychosocial stress (i.e., moving to a new housing environment; Move, n = 8), moderate energy imbalance (Exercise + Diet, n = 9); and all stressors in combination (Move + Exercise + Diet, n = 10). Food intake, body weight, menstrual cyclicity, and reproductive hormones were assessed for two control menstrual cycles followed by two experimental cycles during which the monkeys experienced the stressors. Abnormal cycles were considered to be abnormally long or anovulatory cycles. Few abnormal cycles occurred in the Move group (1 of 8 monkeys) and in the Exercise + Diet group (1 of 9 monkeys). In contrast, 7 of 10 monkeys in the Move + Exercise + Diet group displayed at least one abnormal cycle (chi(2) = 9.61, P = 0.008). These findings suggest that infertility due to hypothalamic hypogonadism can result from the combination of commonplace, seemingly minor stressors that often escape clinical attention.