Prostaglandin E2 Increases Lentiviral Vector Transduction Efficiency of Adult Human Hematopoietic Stem and Progenitor Cells.

Gene therapy currently in development for hemoglobinopathies utilizes ex vivo lentiviral transduction of CD34+ hematopoietic stem and progenitor cells (HSPCs). A small-molecule screen identified prostaglandin E2 (PGE2) as a positive mediator of lentiviral transduction of CD34+ cells. Supplementation with PGE2 increased lentiviral vector (LVV) transduction of CD34+ cells approximately 2-fold compared to control transduction methods with no effect on cell viability. Transduction efficiency was consistently increased in primary CD34+ cells from multiple normal human donors and from patients with ß-thalassemia or sickle cell disease. Notably, PGE2 increased transduction of repopulating human HSPCs in an immune-deficient (nonobese diabetic/severe combined immunodeficiency/interleukin-2 gamma receptor null [NSG]) xenotransplantation mouse model without evidence of in vivo toxicity, lineage bias, or a de novo bias of lentiviral integration sites. These data suggest that PGE2 improves lentiviral transduction and increases vector copy number, therefore resulting in increased transgene expression. As a result, PGE2 may be useful in clinical gene therapy applications using lentivirally modified HSPCs.

Responses to the Human Intruder Test are related to hair cortisol phenotype and sex in rhesus macaques (Macaca mulatta).

Measurement of cortisol in hair provides a chronic index of hypothalamic-pituitary-adrenal (HPA) axis activity and has been applied to assessments of temperament (stable behavioral differences between individuals). However, the extent to which chronically high HPA axis activity relates to a correspondingly high degree of behavioral reactivity is as yet unknown. Therefore, the goal of the present experiment was to assess the relationship between hair cortisol and a reactive temperament. We administered the Human Intruder Test (HIT) twice to 145 (80 male) rhesus macaques (Macaca mulatta) in order to assess behavioral reactivity. The HIT presents monkeys with an unfamiliar experimenter and is composed of a Baseline phase (no intruder) followed by three experimental phases in which the orientation of the intruder changes (Profile, Stare, Back). Behavioral responses to the test were videotaped and behaviors thought to reflect a reactive response to the intruder were scored for duration. Hair samples collected within ±1 month of the first HIT session were analyzed for cortisol by enzyme immunoassay. Subjects were assigned to three groups based on hair cortisol concentration: high, intermediate, and low cortisol phenotypes. Monkeys with the high cortisol phenotype were more reactive to the presence of the intruder than those with the low cortisol phenotype: they were more aggressive, scratched more, and spent more time in the back half of the cage. Males yawned significantly more while females spent more time immobile and in the back of the cage. Overall, monkeys with higher hair cortisol demonstrated an exaggerated response to the presence of the human intruder, supporting a relationship between high levels of chronic HPA axis activity and a reactive temperament. These results indicate that high levels of HPA axis activity, which may result from either genetic variation or environmental stress, correspond with heightened behavioral responses to a stressful experience. Am. J. Primatol. 79:e22526, 2017. © 2016 Wiley Periodicals, Inc.

Inhaled oxytocin increases positive social behaviors in newborn macaques.

Early caregiver-infant interactions are critical for infants' socioemotional and cognitive development. Several hormones and neuromodulators, including oxytocin, affect these interactions. Exogenous oxytocin promotes social behaviors in several species, including human and nonhuman primates. Although exogenous oxytocin increases social function in adults--including expression recognition and affiliation--it is unknown whether oxytocin can increase social interactions in infants. We hypothesized that nebulized oxytocin would increase affiliative social behaviors and such effects would be modulated by infants' social skills, measured earlier in development. We also hypothesized that oxytocin's effects on social behaviors may be due to its anxiolytic effects. We tested these hypotheses in a blind study by nebulizing 7- to 14-d-old macaques (n = 28) with oxytocin or saline. Following oxytocin administration, infants' facial gesturing at a human caregiver increased, and infants' salivary oxytocin was positively correlated with the time spent in close proximity to a caregiver. Infants' imitative skill (measured earlier in development: 1-7 d of age) predicted oxytocin-associated increases in affiliative behaviors--lip smacking, visual attention to a caregiver, and time in close proximity to a caregiver--suggesting that infants with higher propensities for positive social interactions are more sensitive to exogenous oxytocin. Oxytocin also decreased salivary cortisol, but not stress-related behaviors (e.g., scratching), suggesting the possibility of some anxiolytic effects. To our knowledge, this study provides the first evidence that oxytocin increases positive social behaviors in newborns. This information is of critical importance for potential interventions aimed at ameliorating inadequate social behaviors in infants with higher likelihood of developing neurodevelopmental disorder.

Rhesus macaques (Macaca mulatta) displaying self-injurious behavior show more sleep disruption than controls.

Self-injurious behavior (SIB) is a pathology observed in both humans and animals. In humans, SIB has been linked to various mental health conditions that are also associated with significant sleep disruption. In rhesus macaques, SIB consists of self-directed biting which can range from mild skin abrasions to wounds requiring veterinary care. However, only one study suggests possible sleep disruption in macaques with SIB. We evaluated sleep disruption using a noninvasive system (infra-red camera and a video surveillance program) which created videos for every movement over the nighttime hours. Nighttime activity was examined in 13 macaques (three females) of which six were classified as having SIB (one female). Each monkey was studied for a total of 6 nights spanning a period of 4 months. Measures included total movement time (TMT), time moving in the first hour (HR1), time moving in the last hour (HR11), and number of videos <10 secs, ≥10 secs, and ≥30 secs in length. Overall, SIB monkeys had higher TMT (p < 0.01), higher HR1 (p<0.001), and generated more videos ≥10 secs (p < 0.01) and ≥30 secs (p < 0.01). Thus, SIB monkeys showed significant sleep disruption. A four-fold difference between SIB and control monkeys in the ≥30 secs videos revealed many more significant awakenings in the SIB group. Overall higher nighttime activity, in the first hour but not in the last hour, is consistent with sleep-onset insomnia in humans. Whether increased nighttime activity contributes to the SIB condition during the day or, conversely, SIB causes higher nighttime activity remains undetermined.

Stress, the HPA axis, and nonhuman primate well-being: A review.

Numerous stressors are routinely encountered by wild-living primates (e.g., food scarcity, predation, aggressive interactions, and parasitism). Although many of these stressors are eliminated in laboratory environments, other stressors may be present in that access to space and social partners is often restricted. Stress affects many physiological systems including the hypothalamic-pituitary-adrenocortical (HPA) axis, which is the focus of this review. The glucocorticoid, cortisol, is the ultimate output of this system in nonhuman primates, and levels of this hormone are used as an index of stress. Researchers can measure cortisol from several sampling matrices that include blood, saliva, urine, faeces, and hair. A comparison of the advantages and disadvantages of each sampling matrix is provided to aid researchers in selecting an optimal strategy for their research. Stress and its relationship to welfare have been examined in nonhuman primates using two complimentary approaches: comparing baseline cortisol levels under different conditions, or determining the reactivity of the system through exposure to a stressor. Much of this work is focused on colony management practices and developmental models of abnormal behaviour. Certain colony practices are known to increase stress at least temporarily. Both blood sampling and relocation are examples of this effect, and efforts have been made to reduce some of the more stressful aspects of these procedures. In contrast, other colony management practices such as social housing and environmental enrichment are hypothesized to reduce stress. Testing this hypothesis by comparing baseline cortisol levels has not proved useful, probably due to "floor" effects; however, social buffering studies have shown the powerful role of social housing in mitigating reactions of nonhuman primates to stressful events. Models of abnormal behaviour come from two sources: experimentally induced alterations in early experience (e.g., nursery rearing), and the spontaneous development of behavioural pathology (e.g., self-injurious behaviour). Investigators have often assumed that abnormal behaviour is a marker for stress and thus such monkeys are predicted to have higher cortisol levels than controls. However, an emerging finding is that monkeys with abnormal behaviour are more likely to show a pattern of lowered cortisol concentrations which may reflect either an altered set point or a blunting of the stress response system. These findings parallel human clinical studies demonstrating that neuropsychiatric disorders may be associated with either increased or decreased activity of the HPA system, depending on the aetiology and manifestation of the disorder and their potential influence in provoking allostatic shifts in system functioning.

Integrating Geriatrics Knowledge into a Medical Student Clerkship Using Twitter Poll.

A controlled, prospective, 2-year cohort observational study was conducted to test whether weekly geriatric questions delivered through Twitter Poll could improve geriatrics knowledge during an internal medicine clerkship for third-year medical students. Pre- and post-rotation test results used a modified University of California, Los Angeles geriatric knowledge test that included questions linked to 26 Association of American Medical Colleges geriatric competencies for medical students. Data were analyzed using a general linear model repeated-measure design and Student t-test. The primary outcome showed that Twitter Poll participants had more than twice the geriatrics knowledge (p = .002) than students who did not use Twitter Poll. Subset analysis showed different test performances according to sex (p = .03), training site (p = .002), and cohort (p = .003). This study is the first demonstration of Twitter Poll efficacy in medical education and raises questions about whether it could be even more effective if linked to spaced timing of didactic content or supported by annotated answers to geriatrics questions. J Am Geriatr Soc 66:2389-2393, 2018.

Relationships between affiliative social behavior and hair cortisol concentrations in semi-free ranging rhesus monkeys.

Sociality is a fundamental aspect of human behavior and health. One benefit of affiliative social relationships is reduced short-term levels of glucocorticoids (GCs), which are indicative of physiological stress. Less is known, however, about chronic GC production in relation to affiliative social behavior. To address this issue, we studied a semi-free ranging troop of rhesus macaques (Macaca mulatta) and collected hair samples to measure hair cortisol concentrations (HCCs), as a measure of chronic GC production, during routine biannual exams. We collected social behavior (both aggressive and affiliative) and hair samples for 32 adult female rhesus macaques over one year (Experiment 1). Our results indicated that adult females who initiated higher levels of social affiliation had significantly lower levels of HCCs. Neither the initiation nor the receipt of aggression were significantly related to HCCs in this study. In a second experiment we studied 28 mother-infant dyads for the first 90days postpartum to examine mother-infant facial interactions (i.e. mutual gazing). We analyzed HCCs during weaning approximately one year later, which is a major transitional period. We found that infants that engaged in higher levels of mutual gazing in the first 90days postpartum had significantly lower levels of HCCs during weaning. Finally, we studied 17 infant rhesus macaques (13 males) to examine whether social behavior (such as play) in the first five months of life correlated with infant HCCs over those months (Experiment 3). We found that infant males that engaged in more social play had significantly lower levels of HCCs. By relying on an animal model, our study shows that affiliative social traits are associated with lower long-term GC production. Future research should address the complex interactions between social behavior, chronic GC production, and mental and physical health.

Models of stress in nonhuman primates and their relevance for human psychopathology and endocrine dysfunction.

Stressful life events have been linked to the onset of severe psychopathology and endocrine dysfunction in many patients. Moreover, vulnerability to the later development of such disorders can be increased by stress or adversity during development (e.g., childhood neglect, abuse, or trauma). This review discusses the methodological features and results of various models of stress in nonhuman primates in the context of their potential relevance for human psychopathology and endocrine dysfunction, particularly mood disorders and dysregulation of the hypothalamic-pituitary-adrenocortical (HPA) system. Such models have typically examined the effects of stress on the animals' behavior, endocrine function (primarily the HPA and hypothalamic-pituitary-gonadal systems), and, in some cases, immune status. Manipulations such as relocation and/or removal of an animal from its current social group or, alternatively, formation of a new social group can have adverse effects on all of these outcome measures that may be either transient or more persistent depending on the species, sex, and other experimental conditions. Social primates may also experience significant stress associated with their rank in the group's dominance hierarchy. Finally, stress during prenatal development or during the early postnatal period may have long-lasting neurobiological and endocrine effects that manifest in an altered ability to cope behaviorally and physiologically with later challenges. Whereas early exposure to severe stress usually results in deficient coping abilities, certain kinds of milder stressors can promote subsequent resilience in the animal. We conclude that studies of stress in nonhuman primates can model many features of stress exposure in human populations and that such studies can play a valuable role in helping to elucidate the mechanisms underlying the role of stress in human psychopathology and endocrine dysfunction.

Extraction and analysis of cortisol from human and monkey hair.

The stress hormone cortisol (CORT) is slowly incorporated into the growing hair shaft of humans, nonhuman primates, and other mammals. We developed and validated a method for CORT extraction and analysis from rhesus monkey hair and subsequently adapted this method for use with human scalp hair. In contrast to CORT "point samples" obtained from plasma or saliva, hair CORT provides an integrated measure of hypothalamic-pituitary-adrenocortical (HPA) system activity, and thus physiological stress, during the period of hormone incorporation. Because human scalp hair grows at an average rate of 1 cm/month, CORT levels obtained from hair segments several cm in length can potentially serve as a biomarker of stress experienced over a number of months. In our method, each hair sample is first washed twice in isopropanol to remove any CORT from the outside of the hair shaft that has been deposited from sweat or sebum. After drying, the sample is ground to a fine powder to break up the hair's protein matrix and increase the surface area for extraction. CORT from the interior of the hair shaft is extracted into methanol, the methanol is evaporated, and the extract is reconstituted in assay buffer. Extracted CORT, along with standards and quality controls, is then analyzed by means of a sensitive and specific commercially available enzyme immunoassay (EIA) kit. Readout from the EIA is converted to pg CORT per mg powdered hair weight. This method has been used in our laboratory to analyze hair CORT in humans, several species of macaque monkeys, marmosets, dogs, and polar bears. Many studies both from our lab and from other research groups have demonstrated the broad applicability of hair CORT for assessing chronic stress exposure in natural as well as laboratory settings.

Hair loss and hypothalamic-pituitary-adrenocortical axis activity in captive rhesus macaques (Macaca mulatta).

Hair loss is a common problem in captive macaque colonies. A potential factor is the possible influence of stressful environments in the development of hair loss. We examined the relationship between hair loss and chronic hypothalamic-pituitary-adrenal (HPA) axis activity by measuring cortisol in hair. Adult male and female rhesus macaques housed at 3 primate facilities in the United States were screened for degree of hair loss and observed for evidence of hair-plucking behavior. Hair samples and photographic data were obtained from 99 subjects, none of which were hair-pluckers. Macaques with greater than 30% hair loss (alopecia group) showed higher concentrations of hair cortisol than did those with less than 5% hair loss (control group), a finding that was unrelated to age, body weight, or the month in which the sample was collected. Hair loss scores were positively correlated with hair cortisol levels across all monkeys and within the alopecic group alone. In addition, the strong relationship between hair cortisol and alopecia was noted in 2 but not the third facility. Friction with cage surfaces appeared to contribute to hair loss in 18 monkeys. These findings suggest that stress may be one of several factors related to hair loss in some captive nonhuman primates, although whether this relationship is causal or merely correlational is unclear. Moreover, the source of the additional cortisol in the hair of alopecic monkeys (that is, from the circulation or from local synthesis in the skin) remains to be determined.

Effects of shampoo and water washing on hair cortisol concentrations.

BACKGROUND: Measurement of cortisol in hair is an emerging biomarker for chronic stress in human and nonhuman primates. Currently unknown, however, is the extent of potential cortisol loss from hair that has been repeatedly exposed to shampoo and/or water. METHODS: Pooled hair samples from 20 rhesus monkeys were subjected to five treatment conditions: 10, 20, or 30 shampoo washes, 20 water-only washes, or a no-wash control. For each wash, hair was exposed to a dilute shampoo solution or tap water for 45 s, rinsed 4 times with tap water, and rapidly dried. Samples were then processed for cortisol extraction and analysis using previously published methods. RESULTS: Hair cortisol levels were significantly reduced by washing, with an inverse relationship between number of shampoo washes and the cortisol concentration. This effect was mainly due to water exposure, as cortisol levels following 20 water-only washes were similar to those following 20 shampoo treatments. CONCLUSIONS: Repeated exposure to water with or without shampoo appears to leach cortisol from hair, yielding values that underestimate the amount of chronic hormone deposition within the shaft. Collecting samples proximal to the scalp and obtaining hair washing frequency data may be valuable when conducting human hair cortisol studies.