Female Macaques Research Articles

Acute Radiation Effects, the H-ARS in the Non-human Primate: A Review and New Data for the Cynomolgus Macaque with Reference to the Rhesus Macaque.

Medical countermeasure development under the US Food and Drug Administration animal rule requires validated animal models of acute radiation effects. The key large animal model is the non-human primate, rhesus macaque. To date, only the rhesus macaque has been used for both critical supportive data and pivotal efficacy trials seeking US Food and Drug Administration approval. The potential for use of the rhesus for other high priority studies such as vaccine development underscores the need to identify another non-human primate model to account for the current lack of rhesus for medical countermeasure development. The cynomolgus macaque, Macaca fascicularis, has an existing database of medical countermeasure development against the hematopoietic acute radiation syndrome, as well as the use of radiation exposure protocols that mimic the likely nonuniform and heterogenous exposure consequent to a nuclear terrorist event. The review herein describes published studies of adult male cynomolgus macaques that used two exposure protocols-unilateral, nonuniform total-body irradiation and partial-body irradiation with bone marrow sparing-with the administration of subject-based medical management to assess mitigation against the hematopoietic acute radiation syndrome. These studies assessed the efficacy of cytokine combinations and cell-based therapy to mitigate acute radiation-induced myelosuppression. Both therapeutics were shown to mitigate the myelosuppression of the hematopoietic acute radiation syndrome. Additional studies being presented herein further defined the dose-dependent hematopoietic acute radiation syndrome of cynomolgus and rhesus macaques and a differential dose-dependent effect with young male and female cynomolgus macaques. The database supports the investigation of the cynomolgus macaque as a comparable non-human primate for efficacy testing under the US Food and Drug Administration animal rule. Critical gaps in knowledge required to validate the models and exposure protocols are also identified.

The effects of female-male friendships on male postcopulatory levels of oxytocin and vasopressin, and sperm parameters in Macacaarctoides

Male and female stump-tailed macaques (Macaca arctoides) form close relationships akin to human friendships. Oxytocin and vasopressin modulate these and other social relationships and reproductive behavior and physiology in various mammal species. Besides the behavioral effects of oxytocin, this hormone plays an essential role in the ejaculatory process, favoring sperm transport upward the female reproductive tract. Therefore, we investigated the influence of friendships on postcopulatory serum levels of oxytocin and vasopressin in the stump-tailed macaque (Macaca arctoides). In addition, we searched for a correlation between this kind of social relationship and sperm transport in the vagina during the periovulatory and luteal phases. Six female and six male adult macaques having different friendship indices served as experimental animals. Allocated in 57 mating pairs combinations, these animals were allowed to copulate once in the luteal and periovulatory phases. Blood samples were collected from each animal finishing copulation to measure oxytocin and vasopressin. Afterward, we profoundly sedated the females and collected three semen samples from the vagina every 10 min to perform spermatobioscopies. Males' post-copulation oxytocin values increased along with the friendship index, while vasopressin behaves oppositely. Sperm concentration and immotile and motile sperm decreased from one sample to another as male-female closeness increased. Finally, in the periovulatory phase, only in the first vaginal sample, sperm velocities significantly increased with friendship indices. Our results showed that in stump-tailed macaques, heterosexual friendships promote higher postcopulatory oxytocin concentrations and better physiological conditions to males, which probably enhance reproductive success.

Retrospective Evaluation of a Minor Dietary Change in Non-Diabetic Group-Housed Long-Tailed Macaques (Macaca fascicularis).

Simple SummaryMacaques in captivity are prone to becoming overweight and obese, which may cause several health and welfare problems. Diet likely plays an important role herein. In an attempt to reduce overweight incidence and related health problems, a minor dietary change was implemented in our long-tailed macaque breeding colony. The provisioning of bread was replaced by grains and vegetables, while the basic diet of monkey chow remained the same. Overweight status did not differ after dietary change, but some biochemical parameters related to glycemic response and lipid metabolism improved. This study emphasizes the importance of evaluating husbandry changes and shows that relatively minor dietary adjustments may improve animal health and welfare.Macaques in captivity are prone to becoming overweight and obese, which may cause several health problems. A diet that mimics the natural diet of macaques may prevent these problems and improve animal welfare. Adjusting captive diets towards a more natural composition may include increasing fiber content and lowering the glycemic index, i.e., reducing the impact on blood glucose levels. Such a dietary change was implemented in our long-tailed macaque (Macaca fascicularis) breeding colony. The basic diet of monkey chow pellets remained the same, while the supplementary provisioning of bread was replaced by grains and vegetables. This study is a retrospective evaluation, based on electronic health records, that investigated whether this minor dietary change had a beneficial effect on relative adiposity and overweight-related health parameters in 44 non-diabetic, group-housed, female long-tailed macaques. Relative adiposity was measured with a weight-for-height index and blood samples were collected during yearly health checks. Glycemic response and lipid metabolism were evaluated using several biochemical parameters. Relative adiposity and overweight status did not differ after dietary change. Yet, relatively heavy individuals generally lost body weight, while relatively lean individuals gained body weight, leading to a more balanced body weight dynamic. Dietary change did not affect HbA1c and triglyceride levels, while fructosamine and cholesterol levels were significantly reduced. Thus, the minor dietary change had no significant effect on overweight status, but some biochemical parameters related to the risk of diabetes and cardiovascular disease were positively affected. This study emphasizes the importance of evaluating husbandry changes and that critically reviewing husbandry practices can provide valuable insights to improve animal health and welfare.

Chronic binge alcohol and ovariectomy-mediated impaired insulin responsiveness in SIV-infected female rhesus macaques.

Aging people living with HIV (PLWH), especially postmenopausal women may be at higher risk of comorbidities associated with HIV, antiretroviral therapy (ART), hypogonadism, and at-risk alcohol use. Our studies in simian immunodeficiency virus (SIV)-infected male macaques demonstrated that chronic binge alcohol (CBA) reduced acute insulin response to glucose (AIRG), and at-risk alcohol use decreased HOMA-β in PLWH. The objective of this study was to examine the impact of ovariectomy (OVX) on glucose-insulin dynamics and integrity of pancreatic endocrine function in CBA/SIV-infected female macaques. Female macaques were administered CBA (12-15 g/kg/wk) or isovolumetric water (VEH) intragastrically. Three months after initiation of CBA/VEH administration, all macaques were infected with SIVmac251, and initiated on antiretroviral therapy (ART) 2.5 mo postinfection. After 1 mo of ART, macaques were randomized to OVX or sham surgeries (n = 7 or 8/group), and euthanized 8 mo post-OVX (study endpoint). Frequently sampled intravenous glucose tolerance tests (FSIVGTT) were performed at selected time points. Pancreatic gene expression and islet morphology were determined at study endpoint. There was a main effect of CBA to decrease AIRG at Pre-SIV and study endpoint. There were no statistically significant OVX effects on AIRG (P = 0.06). CBA and OVX decreased the expression of pancreatic markers of insulin docking and release. OVX increased endoplasmic stress markers. CBA but not OVX impaired glucose-insulin expression dynamics in SIV-infected female macaques. Both CBA and OVX altered integrity of pancreatic endocrine function. These findings suggest increased vulnerability of PLWH to overt metabolic dysfunction that may be exacerbated by alcohol use and ovarian hormone loss.

Tubectomy of Pregnant and Non-pregnant Female Balinese Macaques (Macaca Fascicularis) With Post-operative Monitoring.

Worldwide, primates, and humans increasingly share habitats and often enter in conflict when primates thrive in human-dominated environments, calling for special management measures. Reproductive control is increasingly used to manage population growth but very few monitoring data are available. Therefore, the efficiency and implications of such programs require a careful examination. In the context of a contraception program in wild female long-tailed macaques in Ubud, Bali, conducted over four successive campaigns between 2017 and 2019, including 140 females (i.e., 41.9% of the reproductive females of the population in 2019), modifications of an endoscopic tubectomy procedure, a permanent sterilization method, clinical evaluation of this method, and the post-operative monitoring results of the neutered females after release are described. This surgical approach was applicable for pregnant females: 28.6% of the treated females were pregnant at the time of the surgery. The procedure used a single lateral port to reach and cauterize both oviducts in non-pregnant as well as in early to mid-term pregnant females. Pregnant females nearer to term required a second lateral port to access both oviducts masked by the size of the gravid uterus. Moreover, bipolar thermocauterization was utilized successfully without resection to realize the tubectomy. The average duration of the laparoscopic surgery was 14 min for non-pregnant females and 22 min for pregnant females. Animals were released 3 h 22 min in average following their capture. This short holding time, recommended for free-ranging primates, was made possible by the minimal invasiveness of the sterilization approach. A laparoscopic post-operative evaluation conducted on two patients during the following campaign confirmed that the oviducts were definitely disrupted and no longer patent. Moreover, no new pregnancies in sterilized females were recorded during the 3-year observation period. The survival rate of the treated females 6 months after sterilization was high (96.3%) with no major post-operative complications clinically recorded. Among females that were pregnant during surgery, 81.1% were confirmed to experience term delivery. This study demonstrates the safety and efficiency of endoscopic tubectomy, even for pregnant females, as a mean of wild macaques' population control.

Effect of Infant Presence on Social Networks of Sterilized and Intact Wild Female Balinese Macaques (Macaca fascicularis).

Simple SummaryIn primates, social interactions are significantly influenced by the female reproductive cycle and the presence of infants. In particular, unweaned infants act as amplifiers of social interactions: they facilitate contact between group members while also triggering occasional conflicts. Sterilized females no longer having young offspring could see their social integration modified. We studied intact and recently sterilized wild female long-tailed macaques (Macaca fascicularis) to see if infant presence was influencing the females’ place and role in the social group. We used the social network analysis tool to compare grooming and proximity to other females for females in three nursing conditions (with young infant (YI), with old infant (OI), and non-nursing (NN). YI females were less involved in grooming but stayed in closer proximity to other females than OI and NN females. We suggest that YI females keeping proximity to others was a way to maximize their infants’ protection, while avoiding too direct social interactions, such as grooming, to protect them from kidnapping risk. Overall, sterilization did not deteriorate female social integration, at least shortly after the surgery. Further research should track the social status of non-nursing females over a long time period, in aid of making sound population management decisions.Contraception is increasingly used to control wild animal populations. However, as reproductive condition influences social interactions in primates, the absence of new offspring could influence the females’ social integration. We studied two groups of wild macaques (Macaca fascicularis) including females recently sterilized in the Ubud Monkey Forest, Indonesia. We used social network analysis to examine female grooming and proximity networks and investigated the role of infant presence on social centrality and group connectivity, while controlling for the fertility status (sterilized N = 14, intact N = 34). We compared the ego networks of females experiencing different nursing conditions (young infant (YI) vs. old infant (OI) vs. non-nursing (NN) females). YI females were less central in the grooming network than other females while being more central in proximity networks, suggesting they could keep proximity within the group to protect their infant from hazards, while decreasing direct grooming interactions, involving potential risks such as kidnapping. The centrality of sterilized and intact females was similar, except for the proximity network where sterilized females had more partners and a better group connectivity. These results confirm the influence of nursing condition in female macaque social networks and did not show any negative short-term effects of sterilization on social integration.

Investigating the role of endogenous estrogens, hormone replacement therapy, and blockade of estrogen receptor-\u03b1 activity on breast metabolic signaling

PurposeMenopause is associated with an increased risk of estrogen receptor-positive (ER +) breast cancer. To characterize the metabolic shifts associated with reduced estrogen bioavailability on breast tissue, metabolomics was performed from ovary-intact and ovariectomized (OVX) female non-human primates (NHP). The effects of exogenous estrogen administration or estrogen receptor blockade (tamoxifen treatment) on menopause-induced metabolic changes were also investigated.MethodsBilateral ovariectomies were performed on female cynomolgus macaques (Macaca fascicularis) to model menopause. OVX NHP were then divided into untreated (n = 13), conjugated equine estrogen (CEE)-treated (n= 13), or tamoxifen-treated (n = 13) subgroups and followed for 3 years. Aged-matched ovary-intact female NHP (n = 12) were used as a premenopausal comparison group. Metabolomics was performed on snap-frozen breast tissue.ResultsChanges in several different metabolic biochemicals were noted, particularly in glucose and fatty acid metabolism. Specifically, glycolytic, Krebs cycle, acylcarnitines, and phospholipid metabolites were elevated in breast tissue from ovary-intact NHP and OVX + CEE in relation to the OVX and OVX + tamoxifen group. In contrast, treatment with CEE and tamoxifen decreased several cholesterol metabolites, compared to the ovary-intact and OVX NHP. These changes were accompanied by elevated bile acid metabolites in the ovary-intact group.ConclusionAlterations in estrogen bioavailability are associated with changes in the mammary tissue metabolome, particularly in glucose and fatty acid metabolism. Changes in these pathways may represent a bioenergetic shift in gland metabolism at menopause that may affect breast cancer risk.

The effect of reproductive state on activity budget, feeding behavior, and urinary C-peptide levels in wild female Assamese macaques

The source of maternal energy supporting reproduction (i.e., stored or incoming) is an important factor determining different breeding strategies (capital, income or mixed) in female mammals. Key periods of energy storage and allocation might induce behavioral and physiological shifts in females, and investigating their distribution throughout reproduction helps in determining vulnerable phases shaping female reproductive success. Here, we examined the effects of reproductive state on activity budget, feeding behavior, and urinary C-peptide (uCP) levels, a physiological marker of energy balance, in 43 wild female Assamese macaques (Macaca assamensis). Over a 13-month study period, we collected 96,266 instantaneous records of activity and 905 urine samples. We found that early lactating females and non-gestating–non-lactating females follow an energy-saving strategy consisting of resting more at the expense of feeding and consuming mostly fruits which contributed to enhancing their energy intake and feeding efficiency. We found an opposite pattern in gestating and late lactating females who feed more at the expense of resting and consume mostly seeds, providing a fiber-rich diet. Storing food into cheek pouches increased throughout gestation while it decreased all along with lactation. Lastly, we found the highest uCP levels during late gestation. Our results reflect different feeding adaptations in response to the energetic costs of reproduction and suggest a critical role of fat accumulation before conception and metabolizing fat during gestation and lactation. Overall, our study provides an integrative picture of the energetics of reproduction in a seasonal species and contributes to our understanding of the diversity of behavioral and physiological adaptations shaping female reproductive success.Significance statementTo offset their substantial energetic investment in reproduction, mammalian females may modify their behavior and the way they extract energy from their environment. In addition, as a result of heightened energy expenditure, female reproduction might trigger physiological shifts. To date, most studies investigated the energetic costs of female reproduction using either a behavioral or a physiological approach. To arrive at a more comprehensive picture, we combined behavioral data with a physiological marker of energy balance, i.e., urinary C-peptide, in a seasonal primate species in its natural habitat. Our results indicate that throughout the reproductive cycle, behavioral and physiological adaptations operate concomitantly, inducing modifications in female activity budget, feeding behavior, and suggesting shifts in fat use. Overall, our results illustrate the relevance of combining data on behavior and hormones to investigate breeding strategies in coping with the energetic costs of reproduction.

Temperament Predicts the Quality of Social Interactions in Captive Female Rhesus Macaques (Macaca mulatta).

Simple SummaryRhesus macaques are highly social animals that are used extensively in research. Providing them with a supportive social environment by pairing individuals with compatible partners improves their life quality and also their use as research animals. Therefore, identifying suitable social partners will benefit both the animals and the science. Previous work showed that female rhesus macaques were more likely to form successful pairs that exhibit little aggression if they showed similarities in certain personality traits measured in infancy. In the current study, we assessed the relationships between the same personality traits and the quality of social interactions between partners. This is important since by ensuring that animals benefit the most from being paired to a compatible partner, we are more likely to expose the animals to positive experiences instead of simply avoiding negative ones. We found that pairs with similar emotionality scores showed more positive social interactions, and pairs with similar nervous temperament used fewer behavioral signals to communicate their dominance relationship. Moreover, pairs that paid more attention to unfamiliar monkey faces were more anxious. These results highlight the importance of taking the animals’ personalities into consideration when attempting to match social partners and provide them with an environment that fits their needs.Previous reports suggest that female macaques with greater similarity in emotionality and nervous temperament, as evaluated in a well-established BioBehavioral Assessment (BBA) at the California National Primate Research Center, were more likely to form successful pairs. We tested whether the same measures can also predict the quality of social interactions among 20 female rhesus macaque pairs. We correlated the pairs’ emotionality and nervous temperament scores obtained in infancy and the levels of behaviors recorded systematically during the pairing process years later. Supporting previous findings, partners with similar emotionality scores were more affiliative, and pairs with similar nervous temperament expressed less dominance/submissive behavior. Exploratorily, we found that pairs that were better at processing social information (part of BBA) were also more anxious. Such animals should be prioritized to be introduced in rooms that house calmer, less aggressive animals and provide opportunities for hiding to alleviate their anxiety. Indeed, positive social experiences not only promote animal welfare, but also reduce stress related confounds and unexplained data variability. Therefore, by incorporating the animals’ temperament into the pair configuration process we increase the likelihood of forming high-quality pairs, both in terms of welfare and the research of which they are a part.

Transcriptional, Epigenetic, and Functional Reprogramming of Monocytes From Non-Human Primates Following Chronic Alcohol Drinking.

Chronic heavy drinking (CHD) of alcohol is a known risk factor for increased susceptibility to bacterial and viral infection as well as impaired wound healing. Evidence suggests that these defects are mediated by a dysregulated inflammatory response originating from myeloid cells, notably monocytes and macrophages, but the mechanisms remain poorly understood. Our ability to study CHD is impacted by the complexities of human drinking patterns and behavior as well as comorbidities and confounding risk factors for patients with alcohol use disorders. To overcome these challenges, we utilized a translational rhesus macaque model of voluntary ethanol self-administration that closely recapitulates human drinking patterns and chronicity. In this study, we examined the effects of CHD on blood monocytes in control and CHD female macaques after 12 months of daily ethanol consumption. While monocytes from CHD female macaques generated a hyper-inflammatory response to ex vivo LPS stimulation, their response to E. coli was dampened. In depth scRNA-Seq analysis of purified monocytes revealed significant shifts in classical monocyte subsets with accumulation of cells expressing markers of hypoxia (HIF1A) and inflammation (NFkB signaling pathway) in CHD macaques. The increased presence of monocyte subsets skewed towards inflammatory phenotypes was complemented by epigenetic analysis, which revealed higher accessibility of promoter regions that regulate genes involved in cytokine signaling pathways. Collectively, data presented in this manuscript demonstrate that CHD shifts classical monocyte subset composition and primes the monocytes towards a more hyper-inflammatory response to LPS, but compromised pathogen response.

Ageing and chronic disease-related changes in the morphometric characteristics of ovarian follicles in cynomolgus monkeys (Macaca fascicularis).

How is the localisation of ovarian follicles affected by ageing and chronic diseases? Ovarian follicles shift deeper towards the medulla, due to thickening of the tunica albuginea (TA), with ageing and some major common chronic diseases. The ovary undergoes morphological and functional changes with ageing. The follicular pool follows these changes with alterations in the amount and distribution of residual follicles. Diseases causing a chronic inflammatory process are associated with morphological changes and impaired ovarian function. We conducted a cross-sectional study, examining 90 ovaries from 90 female monkeys. The samples were collected from April 2018 to March 2019 at Tsukuba Primate Research Center in National Institutes of Biomedical Innovation, Health and Nutrition, Japan. Ovarian samples were obtained from cynomolgus monkeys that died from natural causes or were euthanised. Ovarian sections were stained with haematoxylin and eosin (H&E) for histological analyses. In ovarian sections from 64 female macaques aged 0-25 years, a total of 13 743 follicles at different developmental stages (primordial, intermediary, primary, early secondary and late secondary) were assessed to determine the depth of each follicle from the outer surface of the ovarian cortex to the far end of the follicle, by using a digital imaging software. TA thickness was measured as sum of basal membrane and tunica collagen layer for each ovary under H&E staining. To explore the possibility of age-related trends in ovarian morphometric characteristics, samples were divided into four different age groups (0-3 years (pre-menarche), 4-9 years, 10-14 years and 15-20 years). To evaluate the effect of common chronic diseases on ovarian morphometric characteristics, macaques with diabetes mellitus (DM) (n = 10), endometriosis (n = 8) or inflammatory bowel disease (IBD) (n = 8) were compared to age-matched controls without chronic diseases. Ovarian morphometric analysis revealed that the relative location of follicles became deeper in all age groups according to development of follicles (P < 0.05). Total follicle distance from the ovarian surface was increased with ageing (P < 0.05). In a sub-analysis according to developmental stage, only primordial and intermediary follicles were localised deeper with increasing age (P < 0.05). TA thickness was also increased with ageing (P < 0.05). The localisation of the total number of follicles became deeper in ovaries from monkeys with DM, endometriosis or IBD as compared to the control group (P < 0.05). With DM, analysis of follicles distance at almost each developmental stage was significantly deeper compared to controls (P < 0.05) with the exception of early secondary follicles. With endometriosis, follicles at primary and early and late secondary stages were significantly deeper compared to controls (P < 0.05). Also with IBD, follicles at primary and early and late secondary follicles were significantly deeper compared to controls (P < 0.001). The TA was thicker with DM and endometriosis compared to controls (P < 0.05), but not with IBD (P = 0.16). NA. Two-dimensional histology was used to assess follicle localisation. The possibility of minimal variations between the measured distance to the actual distance in a spherical structure cannot be excluded. Additionally, the severity of disease was not assessed. This study is the first step towards enhancing our understanding of how ageing and chronic diseases affect the relative localisation of dormant and developing follicles. These observations, combined with possible future human studies, may have managerial implications in the field of fertility preservation and other conditions involving ovarian tissue cryopreservation. The present work was supported by the Grant-in-Aid for Scientific Research B (19H03801) (to K.K.), Challenging Exploratory Research (18K19624), Japan Agency for Medical Research and Development, Mochida Memorial Foundation for Medical and Pharmaceutical Research, Takeda Science Foundation and Naito Foundation (to K.K.). All authors have no conflicts of interest directly relevant to the content of this article.

Activity synchrony and travel direction synchrony in wild female Japanese macaques

The degree of behavioural synchrony of animals within a group can be considered a reflection of how individuals adjust their behaviours to manage the costs/benefits accompanying group-living. In this study, we focused on activity synchrony and travel direction synchrony as behavioural synchrony. We aimed to quantify the degree of behavioural synchrony and identify which factors can affect the synchrony in wild females of Japanese macaques. Japanese macaques live in female philopatric multi-female and multi-male groups and have a linear dominance hierarchy. The groups are characterized by changing spatio-temporal cohesiveness among group members. Two observers conducted simultaneous focal animal sampling on adult females using global positioning system devices to record locations. The overall degree of activity synchrony was positive compared with random, and the degree was highest when macaques were located within visual range of each other. Both activity synchrony and travel direction synchrony were influenced by spatial cohesion, i.e. interindividual distance, which shows that the probabilities of synchrony were higher with individuals located closer. Activity synchrony was also influenced by activity type, showing that the probabilities of synchrony were higher when individuals engaged in foraging. These results suggest that synchronized foraging may be caused by enhanced feeding with other group members when they are closer to each other. Our approach to quantitatively measure spatial dispersal while observing group members simultaneously revealed the roles of spatial cohesion and activity types for determining the degree of behavioural synchrony.

Chronic binge alcohol and ovariectomy dysregulate omental adipose tissue metaboproteome in simian immunodeficiency virus-infected female macaques

Effective antiretroviral therapy (ART) has significantly reduced mortality of people living with HIV (PLWH), and the prevalence of at-risk alcohol use is higher among PLWH. Increased survival and aging of PLWH is associated with increased prevalence of metabolic comorbidities especially among menopausal women, and adipose tissue metabolic dysregulation may be a significant contributing factor. We examined the differential effects of chronic binge alcohol (CBA) administration and ovariectomy (OVX) on the omental adipose tissue (OmAT) proteome in a subset of simian immunodeficiency virus (SIV)-infected macaques of a longitudinal parent study. Quantitative discovery-based proteomics identified 1,429 differentially expressed proteins. Ingenuity Pathway Analysis (IPA) was used to calculate z-scores, or activation predictions, for functional pathways and diseases. Results revealed that protein changes associated with functional pathways centered around the "OmAT metaboproteome profile." Based on z-scores, CBA did not affect functional pathways of metabolic disease but dysregulated proteins involved in adenosine monophosphate-activated protein kinase (AMPK) signaling and lipid metabolism. OVX-mediated proteome changes were predicted to promote pathways involved in glucose- and lipid-associated metabolic disease. Proteins involved in apoptosis, necrosis, and reactive oxygen species (ROS) pathways were also predicted to be activated by OVX and these were predicted to be inhibited by CBA. These results provide evidence for the role of ovarian hormone loss in mediating OmAT metaboproteome dysregulation in SIV and suggest that CBA modifies OVX-associated changes. In the context of OVX, CBA administration produced larger metabolic and cellular effects, which we speculate may reflect a protective role of estrogen against CBA-mediated adipose tissue injury in female SIV-infected macaques.

Social aging in male and female Barbary macaques

Aging brings about notable changes in sociality, with an increasing focus on essential partners in both humans and nonhuman primates. Several studies have shown that older nonhuman primates have fewer social partners and shift their types of interactions. The majority of these studies, however, involved only female individuals. Much less is known about the trajectory of social aging in males. We collected 2180 h of focal observation data in a large age-heterogeneous sample of 34 male and 50 female Barbary macaques (Macaca sylvanus; age range 5-30 years) living in two social groups at the outdoor enclosure at La Forêt des Singes in Rocamadour (France). To track age-related changes in social engagement of both sexes, we used classical behavioral and social network analysis to measure age-related variation in the number of partners, the frequency of affiliative interactions, and the position in the social network (eigenvector centrality and local clustering coefficient). We found that females were more central in most social network metrics than males, that is, had more social partners and were more engaged in affiliative interactions than males. We did not find notable sex differences with age: both sexes showed a similar decline in social activity and energy-demanding activities like jumping or running. Our results thereby extend our knowledge of older nonhuman primates' social life and emphasize that age can have a similar impact on female and male social behavior.

Validation of the Social Responsiveness Scale (SRS) to screen for atypical social behaviors in juvenile macaques.

Primates form strong social bonds and depend on social relationships and networks that provide shared resources and protection critical for survival. Social deficits such as those present in autism spectrum disorder (ASD) and other psychiatric disorders hinder the individual’s functioning in communities. Given that early diagnosis and intervention can improve outcomes and trajectories of ASD, there is a great need for tools to identify early markers for screening/diagnosis, and for translational animal models to uncover biological mechanisms and develop treatments. One of the most widely used screening tools for ASD in children is the Social Responsiveness Scale (SRS), a quantitative measure used to identify individuals with atypical social behaviors. The SRS has been adapted for use in adult rhesus monkeys (Macaca mulatta)–a species very close to humans in terms of social behavior, brain anatomy/connectivity and development–but has not yet been validated or adapted for a necessary downward extension to younger ages matching those for ASD diagnosis in children. The goal of the present study was to adapt and validate the adult macaque SRS (mSRS) in juvenile macaques with age equivalent to mid-childhood in humans. Expert primate coders modified the mSRS to adapt it to rate atypical social behaviors in juvenile macaques living in complex social groups at the Yerkes National Primate Research Center. Construct and face validity of this juvenile mSRS (jmSRS) was determined based on well-established and operationalized measures of social and non-social behaviors in this species using traditional behavioral observations. We found that the jmSRS identifies variability in social responsiveness of juvenile rhesus monkeys and shows strong construct/predictive validity, as well as sensitivity to detect atypical social behaviors in young male and female macaques across social status. Thus, the jmSRS provides a promising tool for translational research on macaque models of children social disorders.

Gut microbiota are associated with sex and age of host: Evidence from semi-provisioned rhesus macaques in southwest Guangxi, China.

Host characteristics, such as sex and age, are closely associated with the structure and function of gut microbiota; however, less is known about the effects of age and sex on the gut microbiota of nonhuman primates, and therefore, our knowledge of interindividual variability in host gut microbiota is limited. In this study, 153 fecal samples from rhesus macaques (Macaca mulatta) were analyzed using high‐throughput 16S rRNA sequencing in order to explore associations between age and sex of the host and their gut microbiota. The results indicated that female macaques had higher alpha diversity and a more unique gut microbiota than did males. The proportion of Proteobacteria, Tenericutes, Cyanobacteria, unclassified bacteria, and Verrucomicrobia was higher in females than that in males. We also found that adults of both sexes had a higher alpha diversity, a higher proportion of norank Ruminococcaceae, Oscillospira, norank Lachnospiraceae, norank Clostridiales, and Succinivibrio, and a lower proportion of Enterococcus than immatures. Functional analyses revealed that the richness of metabolic pathways was higher in females than males and in adults compared with immatures. These results could be attributed to differences in the nutritional requirements and hormone levels of macaques of different sex and age classes. We conclude that variation in the gut microbiota of different sex and age classes of rhesus macaques may be linked to age‐ and sex‐specific differences in nutrient requirements and hormone levels. These results highlight the importance of host age and sex on the structure and function of the gut microbiota and the need to consider physiological traits when conducting studies on the gut microbiota.

High‐Fat High‐Sucrose Diet Increases ACE2 Receptor Expression in Lung and Pancreatic Islets in SIV‐Infected Rhesus Macaques: Implications for Increased Risk for SARS‐CoV‐2 Infection

People with pre‐existing conditions including obesity, diabetes, and kidney disease are at greater risk for complications of COVID‐19, the disease caused by SARS‐CoV‐2 infection. Predisposing risk factors could include poor dietary quality and at‐risk alcohol use. Increased survival and aging of people living with HIV (PLWH) on antiretroviral therapy (ART) is complicated by comorbidities including at‐risk alcohol use, metabolic dysregulation, diabetes, and chronic renal disease, with obesity and metabolic syndrome highest in southern states. We hypothesize that poor dietary quality in combination with at‐risk alcohol use may increase risk factors for SARS‐CoV‐2 infection in PLWH. To test this hypothesis, biospecimen obtained from a longitudinal study in female rhesus macaques (macaca mulatta) were used to determine whether high‐fat high‐sucrose diet (HFD; protein/fat/carbohydrates 16/42/42% of total kcal and 27% sucrose by weight) and chronic binge alcohol administration (CBA) increased expression of the machinery required for SARS‐CoV‐2 cell entry. Female macaques (n=10) were assigned to HFD or standard chow diet (SD; protein/fat/carbohydrates 29/14/57% of total kcal) and CBA (50‐60 mM peak blood alcohol, 5 days/week) or isovolumetric water (VEH) beginning 3 months before SIVmac251 infection. ART was initiated at 2.5 months of SIV infection. Tissue samples including lung, pancreas, and kidney were collected at study endpoint (~12 months post‐SIV infection). Immunohistochemistry was performed on formalin‐fixed, paraffin embedded samples to determine protein expression of angiotensin converting enzyme 2 (ACE2) receptor and transmembrane serine protease 2 (TMPRSS2). Images were obtained at 20x magnification and fluorescence intensity was used for quantification. RNA was extracted from frozen samples and mRNA expression of ACE2 and TMPRSS2 was measured by qPCR and quantified relative to ribosomal protein S13. Normality of data was verified and outliers identified using Grubbs test. Data were then analyzed using a 2 (alcohol) × 2 (diet) ANOVA. Protein expression of ACE2 in the lung (p<0.01, η2=0.59), whole pancreas (p<0.05, η2=0.64), and pancreatic islets (p<0.05, η2=0.44) was greater in HFD‐ than SD‐fed macaques and was not significantly altered by CBA. No alcohol‐ or diet‐mediated differences in ACE2 protein expression were observed for whole kidney or proximal tubules. No alcohol‐ or diet‐mediated differences in protein expression of TMPRSS2 or relative mRNA expression of ACE2 or TMPRSS2 were observed. Results indicate that a high‐fat, high‐sucrose diet increases expression of the ACE2 receptor in SIV‐infected female macaques. Because the ACE2 receptor is required for SARS‐CoV‐2 cell entry, this diet‐mediated increase in expression may increase risk for greater vulnerability to COVID‐19 disease and associated complications. These data provide direct evidence for a link between dietary quality and risk for SARS‐CoV‐2 infection in the context of SIV/HIV infection, urging diet counseling and increased access to higher‐quality foods in this population.

Ovariectomy Increases CD4 + T cell Activation in Simian Immunodeficiency Virus Infection

People living with HIV (PLWH) have a higher prevalence of alcohol use disorder (AUD), which has been associated with accelerated HIV disease progression. Older PLWH, especially postmenopausal women, are at a greater risk for alcohol-related problems. Previous studies have shown that chronic binge alcohol (CBA) administration leads to higher simian immunodeficiency virus (SIV) loads and accelerated progression towards end-stage disease in rhesus macaques. Clinical studies have shown that ovariectomy (OVX) increases T cell activation and serum TNF-alpha levels. We tested the hypothesis that CBA and ovarian hormone loss, resulting from OVX, increases T cell activation in SIV-infected female rhesus macaques. CBA or isovolumetric water was administered through an intragastric catheter for 30 minutes, 5 days a week, with peak blood alcohol concentrations reaching 50-60 mM. After three months of CBA/VEH administration, macaques were infected with SIVMac251 and 2.5 months later initiated on ART. OVX was performed one month after ART initiation, and study endpoint was ~12 months post-SIV infection. Blood samples obtained at study end point were used for flow cytometry analysis with FITC-CD38, PerCP-CD3, PE-CD45, Qdot 655-CD8, ECD-CD20, APC-Cy7-CD4, PE-Cy7-CD16, APC-CD66, Pacific Blue-CD14, and Am Cyan-Live Dead, using a BD Biosciences LSRII flow cytometer and FlowJo software (version 10.7.1). Two-way ANOVA was used to detect significant differences between groups (N = 7-8/group). At study end-point, there was a main effect of OVX to increase activated CD4+CD38+ T cells (p = 0.0171) compared to SHAM. Neither CBA nor OVX had an effect on activation of CD8+ T cells. There was a main effect of OVX (p = 0.0195) and an interaction (p = 0.05) observed for CD4 T cell numbers (cells/ml). Post-hoc analysis showed higher CD4 T cell number in the CBA/SIV/ART/OVX compared to CBA/SIV/ART/SHAM (p = 0.0174) animals. There were no significant differences between the treatment groups for CD20+, CD14+, and CD16+ cells. Our results show that OVX lead to an increase in activation of peripheral CD4+ T cells and an increase in peripheral CD4+ T cells within CBA animals, which could favor a pro-inflammatory environment and an increase in target cells in SIV-infected female macaques.

Chronic binge alcohol impairs glucose‐insulin dynamics in SIV‐infected female rhesus macaques

Background Antiretroviral therapy (ART) has significantly reduced mortality of people living with HIV (PLWH). Older PLWH, especially post-menopausal women, experience complex interactions of HIV, ART, hormonal status, and lifestyle behaviors such as at-risk alcohol use, increasing the risk of metabolic comorbidities. We have previously shown that chronic binge alcohol (CBA) reduces the acute insulin response to a glucose challenge in SIV-infected male macaques and that at-risk alcohol use decreases HOMA-β (a marker for pancreatic beta cell function) in PLWH. Meanwhile, the role of estrogen in pancreatic health maintenance is not fully understood. The objective of this study was to determine the effect of CBA and ovarian hormone loss, resulting from ovariectomy (OVX), on glucose-insulin dynamics and pancreatic integrity in SIV-infected female rhesus macaques. Hypothesis We hypothesized that chronic binge alcohol and ovariectomy would decrease insulin response to a glucose challenge and impair insulin and glucagon expression in pancreatic islets. Methods 28 adult female macaques were used in the study. CBA or isovolumetric water (VEH) was administered through an intragastric catheter for 30 minutes, 5 days a week, with blood alcohol concentrations reaching 50-60 mM. Three months after initiation of CBA/VEH administration, macaques were infected with SIVMac251 and 2.5 months later initiated on ART. After 1 month of ART, macaques underwent sham surgery or ovariectomy. Using a modified frequently sampled intravenous glucose tolerance test (FSIVGTT), blood glucose and serum insulin values were measured. Glucose and insulin dynamic measures including the acute insulin response to glucose (AIRg), insulin sensitivity (Si), disposition index (DI), and glucose effectiveness (Sg) were determined using minimal model software. Insulin and glucagon expression was determined in formalin-fixed, paraffin-embedded pancreatic tissue collected at study endpoint. Corrected total cell fluorescence for insulin and glucagon were calculated using ImageJ software. To confirm these results, total RNA was isolated from pancreas and mRNA expression of insulin and glucagon determined using quantitative PCR. Results There was a main effect of CBA to decrease AIRg (p=0.02) and increase Si (p=0.01). There were no statistically significant effects of OVX on AIRg (p=0.06) or Si (p=0.4). There were also no statistically significant effects of CBA or OVX on DI or Sg. Neither CBA nor OVX altered the protein or mRNA expression of insulin and glucagon in pancreatic islets. Conclusion CBA decreased the acute endogenous insulin release in response to a glucose challenge without significantly altering basal pancreatic islet expression of insulin and glucagon, while OVX did not significantly alter any of these measures. These results indicate that CBA impairs the immediate release of insulin from beta cell insulin secretory vesicles, and we speculate that CBA dysregulates the metabolic signaling pathway of insulin secretion. Determining mechanisms of pancreatic endocrine dysfunction will inform therapeutic strategies to improve alcohol-mediated impaired glucose insulin dynamics in the context of SIV/HIV.

Delay Discounting of Cocaine‐Food Choice in Socially Housed Female and Male Cynomolgus Macaques

At present, there are no FDA-approved pharmacological treatments for cocaine use disorders (CUD). Because there is evidence of sex differences in vulnerability to cocaine abuse (O’Brien and Anthony, 2005) and treatment outcomes (Sugarman et al., 2017), research needs to include both sexes in evaluating treatment strategies. Weafer and de Wit (2014) have suggested that impulsivity is a stronger predictor of drug abuse in females compared to males and lack of inhibitory control may be a mechanism mediating the faster progression of drug use in women compared to men. The overarching goal of the present research is to utilize a “personalized medicine” approach to evaluating the role of environmental and pharmacological interventions on the reinforcing effects of cocaine. In this study, socially housed female (N=5) and male (N=6) cynomolgus monkeys, each implanted with an indwelling intravenous catheter and self-administering cocaine (0.01-0.1 mg/kg/injection) under a concurrent schedule with food (1.0-g banana-flavored pellets) as the alternative, served as subjects. Under these conditions, the frequency of cocaine choice increases in a dose-dependent manner, from <20% choice when 0.01 mg/kg/injection cocaine was available as an alternative to food, to >80% choice when 0.1 mg/kg/injection was the alternative to food; in this preliminary study, no sex or social rank differences were noted. Next, environmental manipulations were studied at both ends of the cocaine dose-response curve in order to assess whether there were sex differences in sensitivity to the reinforcing effects of cocaine. To study impulsivity, we used a delay discounting procedure similar to that described by Woolverton and colleagues (Huskinson et al., 2015, 2016). Differences in sensitivity to changes in cocaine preference when the preferred cocaine dose was available would provide information as to potential sex differences in the reinforcing strength of cocaine. In contrast, when low, non-preferred, cocaine doses were studied and food reinforcement was delayed, sex differences would provide information related to sensitivity of adverse environmental changes on vulnerability to cocaine abuse. In preliminary findings, when delays were imposed on high-dose cocaine, the indifference points (IP; 50% choice between cocaine and food) were not different between males and females (168.88 ± 75.6 seconds vs. 154.94 ± 39.1 seconds, respectively). However, when low-dose cocaine was available and food reinforcement was delayed, males were more sensitive to changes in the environment, as represented by lower IP values (46.74 ± 16.8 seconds vs. 114.39 ± 44.9 seconds, respectively). Studies are currently underway to determine if pharmacological manipulations, with and without delays, would provide additional reductions in cocaine choice, at both ends of the cocaine dose-response curve. An interesting behavioral pharmacology study would involve examine drug effects under conditions in which delayed high-dose cocaine and food-delayed low-dose cocaine produce similar behaviors (i.e., % cocaine choice).