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.     

关于非人灵长类的公正行为的研究

人类关注公正, 非人灵长类也表现出公正行为。本文先以现有研究资料为基础, 以理毛为例分析后认为, 非人灵长类关注投入—收益的对称性, 说明它们可能具备不公正规避这一心理特质; 关于非人灵长类公正行为的实验也表明, 它们不仅比较自身的投入—收益对称性, 而且能在社会比较过程中与其它个体相比。有实验得出期望假设和挫折效应能更好地解释被试的行为, 本文认为, 这些实验结论不一致的主要原因, 是研究者未充分考虑\"投入\"对被试行为的影响。文章在最后进行了总结并提出了三点研究展望。     

Antioxidant treatment in the absence of exogenous lipids and proteins protects rhesus macaque sperm from cryopreservation-induced cell membrane damage

Osmotic stress caused oxidative stress in rhesus macaque sperm, which was alleviated by antioxidant supplementation. The objective of the present study was to demonstrate that cryopreservation of rhesus macaque sperm also induces reactive oxygen species (ROS) production, and to determine whether ROS have an important role in cryopreservation-induced membrane. Additionally, we evaluated the antioxidant capacity of TEST (N-Tris(hydroxymethyl)methyl-2-aminoethanesulfonic acid-Tris) buffer (with 20% egg yolk and 13% skim milk) and supplementation with antioxidants, superoxide dismutase (SOD), catalase (CAT), and α-tocopherol. There was a substantial level of ROS production in both the presence (15% increase in superoxide, P < 0.01; 14% increase in hydrogen peroxide, P < 0.01) and absence of egg yolk (EY) and skim milk (SM; 33% increase in superoxide, P < 0.001; 48% increase in hydrogen peroxide, P < 0.001). Superoxide dismutase provided little membrane protection against ROS, but increased postthaw total and progressive motility by 10% (P < 0.01) and 15% (P < 0.05), respectively. Supplementation with CAT and α-tocopherol in the presence of EY and SM decreased H₂O₂ by 55% (P < 0.01) and 49% (P < 0.001), whereas supplementation with CAT and α-tocopherol in the absence of EY and SM reduced the level of lipid peroxidation by 61% (P < 0.05) and 28% (P < 0.01). In conclusion, this is apparently the first report that cryopreservation of rhesus macaque sperm induced a significant increase in ROS and that antioxidant supplementation (N-Tris(hydroxymethyl)methyl-2-aminoethanesulfonic acid-Tris) can significantly decrease the extent of ROS-induced membrane damage.     

Minimally invasive transabdominal collection of preimplantation embryos from the common marmoset monkey (Callithrix jacchus)

A novel, minimally invasive, transabdominal embryo collection method (transabdominal method) was developed as an alternative to a standard abdominal incision for embryo collection in the common marmoset. The abdominal incision method was used for 304 flushes using 36 female animals, whereas the transabdominal method was used for 488 flushes using 48 females; successful embryo collection rates were 48.0% and 48.4% (P > 0.05), respectively. These techniques were successfully duplicated at another institute (German Primate Center, DPZ). At that institution, successful embryo collection rates were 88.9% and 77.8% for the abdominal incision and transabdominal methods, respectively (P > 0.05), whereas the average numbers of preimplantation embryos obtained per flush were (mean ± SD) 1.91 ± 0.35 and 1.71 ± 0.14 (P > 0.05). The transabdominal method reduced animal stress, did not require incisional wound healing, and enabled successive embryo recoveries to be done much sooner. More embryos in early developmental stages (zygotes/morulae) were recovered using the transabdominal method (76.1%) than the abdominal incision method (52.6%, P < 0.01). In contrast, recovery of arrested or abnormal embryos was not significantly different between these two methods (9.8% and 8.3%). To verify developmental ability of embryos recovered by the transabdominal method, transfer of 28 normal embryos to 14 surrogate mothers yielded a nidation rate of 57%. Five females sustained term pregnancies and eight neonates were born. This novel transabdominal method will facilitate progress in marmoset developmental biology and embryology.     

非人灵长类的杀婴行为及其适应意义

杀婴现象普遍存在于非人灵长类种类中,本文通过综述目前已知的21种非人灵长类的69例杀婴行为记录,探讨了非人灵长类杀婴行为的适应性意义.大多数雄性杀婴行为符合雄性繁殖策略假说,既满足以下3个条件:(1)杀婴雄性与受害婴仔没有父子关系;(2)失去婴猴的母亲会提前进入发情状态;(3)受害雌性随后可能与杀婴雄性交配.不过有些个别的杀婴现象与雄性繁殖利益无关,文中进一步介绍了误伤假说、减少未来竞争者假说、肉食假说和病态行为假说.杀婴无疑会对受害母亲造成巨大的损失.雌性防范杀婴的策略包括直接抵抗、回避接触雄性、积极发情、依靠保护雄性和改变繁殖群组成等多种方式.但是雄性杀婴现象不会完全消失,因为出现该行为的根本原因是雌性防范雄性杀婴的难度较大,而雄性实施杀婴行为的风险较低、利益较高.     

The effect of pseudomonas exotoxin A on cytokine production in whole blood exposed to Pseudomonas aeruginosa

To determine the effect of Pseudomonas aeruginosa exotoxin A (P-ExA) on cytokine production, we studied cytokine release induced by heat-killed P. aeruginosa (HKPA) in human whole blood in the presence or absence of P-ExA. P-ExA (0.01-1 microgram ml(-1)) caused a dose-dependent decrease in HKPA-induced production of tumor necrosis factor alpha (TNF), interleukin (IL-) 10, IL-6 and IL-8 (all P<0.05). P-ExA-induced inhibition of IL-10, IL-6 and IL-8 release was not dependent on reduced TNF concentrations, since the relative attenuation of the production of these cytokines was similar in the presence or absence of a neutralizing anti-TNF antibody. The effect of P-ExA on cytokine production may offer a disadvantage to the host with respect to clearance of the infection.     

Congenital bronchiolo-alveolar airway malformation in a cynomolgus macaque (Macaca fascicularis)

Pulmonary congenital anomalies in animals are rare. Previously reported malformations include accessory lung formation, pulmonary hypoplasia, pulmonary agenesis, and various forms of hamartoma. Congenital bronchiolo-alveolar airway malformation, a new entity, is described in a 1-day-old male cynomolgus macaque. This neonate experienced breathing difficulties shortly after birth and died while therapy was being administered. Grossly, the right lung was markedly increased in size, firm, and pink. Histopathologically, sections of right lung showed irregular bronchiole-like and alveolus-like structures. There was marked widening of alveolar septae by loosely arranged mesenchymal cells and many centrally located capillaries. Alveoli were lined by cuboidal epithelial cells. There were scattered islands of immature cartilage. A grossly enlarged lung containing bronchiole-like and alveolus-like structures, immature cartilage islands, and many capillaries within alveolar septae on histopathologic examination, is inconsistent with previously described congenital pulmonary anomalies in animals and humans.     

Bacteria-phagocyte interactions: emerging tactics in an ancient rivalry

Abstract Although phagocytes appear to have a redundancy of both oxidative and non-oxidative killing mechanisms, nevertheless, bacterial pathogens are still able to evade these defenses in vivo and cause lethal infection. As the mechanisms by which phagocytes function have become detailed at the molecular level, both the recognition of specific bacterial virulence determinants and their effects at specific sites in the phagocytes are also being identified. Knowledge of these interactions may permit the use of immunomodulators either to neutralize these virulence determinants or to enhance the bacterial capabilities of the phagocyte.     

4040 SNPs for genomic analysis in the rhesus macaque (Macaca mulatta)

Although the rhesus macaque (Macaca mulatta) is commonly used for biomedical research and becoming a preferred model for translational medicine, quantification of genome-wide variation has been slow to follow the publication of the genome in 2007. Here we report the properties of 4040 single nucleotide polymorphisms discovered and validated in Chinese and Indian rhesus macaques from captive breeding colonies in the United States. Frequency-matched measures of linkage disequilibrium were much greater in the Indian sample. Although the majority of polymorphisms were shared between the two populations, rare alleles were over twice as common in the Chinese sample. Indian rhesus had higher rates of heterozygosity, as well as previously undetected substructure, potentially due to admixture from Burma in wild populations and demographic events post-captivity.     

Zolpidem Displays Heterogeneity in Its Binding to the Nonhuman Primate Benzodiazepine Receptor In Vivo

Abstract: The distinctive pharmacological activity of zolpidem in rats compared with classical benzodiazepines has been related to its differential affinity for benzodiazepine receptor (BZR) subtypes. By contrast, in nonhuman primates the pharmacological activity of zolpidem was found to be quite similar to that of classical BZR agonists. In an attempt to explain this discrepancy, we examined the ability of zolpidem to differentiate BZR subtypes in vivo in primate brain using positron emission tomography. The BZRs were specifically labeled with [¹¹C]flumazenil. Radiotracer displacement by zolpidem was monophasic in cerebellum and neocortex, with in vivo Hill coefficients close to 1. Conversely, displacement of [¹¹C]flumazenil was biphasic in hippocampus, amygdala, septum, insula, striatum, and pons, with Hill coefficients significantly smaller than 1, suggesting two different binding sites for zolpidem. In these cerebral regions, the half-maximal inhibitory doses for the high-affinity binding site were similar to those found in cerebellum and neocortex and ～100-fold higher for the low-affinity binding site. The low-affinity binding site accounted for <32% of the specific [¹¹C]-flumazenil binding. Such zolpidem binding characteristics contrast with those reported for rodents, where three different binding sites were found. Species differences in binding characteristics may explain why zolpidem has a distinctive pharmacological activity in rodents, whereas its pharmacological activity in primates is quite similar to that of classical BZR agonists, except for the absence of severe effects on memory functions, which may be due to the lack of substantial zolpidem affinity for a distinct BZR subtype in cerebral structures belonging to the limbic system.     

Environmental and social influences on neuroendocrine puberty and behavior in macaques and other nonhuman primates

This article is part of a Special Issue “Puberty and Adolescence”.     

In Vivo High Intrinsic Efficacy of Triazolam: A Positron Emission Tomography Study in Nonhuman Primates

Abstract: The triazolobenzodiazepine triazolam is a central-type benzodiazepine receptor (BZR) ligand that is widely prescribed as a hypnotic agent. Triazolam produces its effects through potentiation of γ-aminobutyric acid-mediated neurotransmission. Findings reported from in vitro binding studies showed some discrepancies concerning the pharmacological characteristics of triazolam. The present study aims to characterize in vivo the biochemical properties of triazolam, i.e., cerebral pharmacokinetics, interaction with BZR, potency, and intrinsic efficacy. Triazolam was studied in living nonhuman primates using positron emission tomography. Two different studies were carried out: (a) a direct study using [¹¹C]triazolam and (b) an indirect competition study using the radiolabeled BZR antagonist [¹¹C]flumazenil. Results showed that, in the brain in vivo, triazolam binds specifically and competitively to the BZR. Its rapid cerebral kinetics is consistent with a hypnotic profile (maximal binding after 23 min, elimination half-life of 202 min). Triazolam is very potent in displacing [¹¹C]flumazenil (ID₅₀= 28 ± 6 μg/kg). Hill analysis of the displacement curve does not show obvious binding-site heterogeneity. Triazolam is 20 times more potent in displacing [¹¹C]flumazenil and 50 times more potent in inhibiting pentylenetetrazol-induced paroxysmal activity than the full benzodiazepine agonist diazepam. Interestingly, the simultaneous use of positron emission tomography and EEG recording allowed us to show that triazolam-positive intrinsic efficacy is slightly higher (20%) than that of diazepam. An attractive hypothesis proposes that the severity of side effects of BZR ligands is proportional to their intrinsic efficacy. Therefore, our study shows that triazolam side effects, as for other benzodiazepines, may be related to its high intrinsic efficacy in vivo.     

Optimized microRNA purification from TRIzol-treated plasma

Background

MicroRNAs (miRNAs) represent new and potentially informative diagnostic targets for disease detection and prognosis. However, little work exists documenting the effect of TRIzol, a common viral inactivation and nucleic acid extraction reagent, on miRNA purification. Here, we developed an optimized protocol for miRNA extraction from plasma samples by evaluating five different RNA extraction kits, TRIzol phase separation, purification additives, and initial plasma sample volume. This method was then used for downstream profiling of plasma miRNAs found in archived samples from one nonhuman primate (NHP) experimentally challenged with Ebola virus by the aerosol route.

Results

Comparison of real-time RT-PCR results for spiked-in and endogenous miRNA sequences determined extraction efficiencies from five different RNA purification kits. These experiments showed that 50 μL plasma processed using the QIAGEN miRNeasy Mini Kit with 5 μg of glycogen as a co-precipitant yielded the highest recovery of endogenous miRNAs. Using this optimized protocol, miRNAs from archived plasma samples of one rhesus macaque challenged with aerosolized Ebola virus was profiled using a targeted real-time PCR array. A total of 519 of the 752 unique miRNAs assayed were present in the plasma samples at day 0 and day 7 (time of death) post-exposure. Statistical analyses revealed 25 sequences significantly up- or down-regulated between day 0 and day 7 post infection, validating the utility of the extraction method for plasma miRNA profiling.

Conclusions

This study contributes to the knowledgebase of circulating miRNA extraction methods and expands on the potential applications of cell-free miRNA profiling for diagnostics and pathogenesis studies. Specifically, we optimized an extraction protocol for miRNAs from TRIzol-inactivated plasma samples that can be used for highly pathogenic viruses.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1299-5) contains supplementary material, which is available to authorized users.     

A high incidence of Shigella-induced arthritis in a primate species: major histocompatibility complex class I molecules associated with resistance and susceptiblity, and their relationship to HLA-B27

 The human major histocompatibility complex (MHC) class I gene, HLA-B27, is a strong risk factor for susceptibility to a group of disorders termed spondyloarthropathies. Rodents that express HLA-B27 develop spondyloarthropathies, implicating HLA-B27 in the etiology of these disorders. To determine whether an HLA-B27-like molecule was associated with spondyloarthropathies in nonhuman primates, we analyzed the MHC class I cDNAs expressed in a cohort of rhesus macaques that developed reactive arthritis after an outbreak of shigellosis. We identified several cDNAs with only limited sequence similarity to HLA-B27. Interestingly, one of these MHC molecules had a B pocket identical to that of HLA-B39. Pool sequencing of radiolabeled peptides bound by this molecule demonstrated that, like HLA-B27 and HLA-B39, it could bind peptides with arginine at the second position. However, extensive analysis of the MHC class I molecules in this cohort revealed no statistically significant association between any particular MHC class I allele and susceptibility to reactive arthritis. Furthermore, none of the rhesus MHC class I molecules bore a strong resemblance to HLA-B27, indicating that reactive arthritis can develop in this animal model in the absence of an HLA-B27-like molecule. Surprisingly, there was a statistically significant association between the rhesus macaque MHC A locus allele, Mamu-A*12, and the absence of reactive arthritis following Shigella infection. Received: 26 July 1999 / Revised: 28 December 1999