Age-related changes in hematological biomarkers in common marmosets.

Researchers and veterinarians often use hematology and clinical chemistry to evaluate animal health. These biomarkers are relatively easy to obtain, and understanding how they change across healthy aging is critical to clinical care and diagnostics for these animals. We aimed to evaluate how clinical biomarkers from a chemistry profile and complete blood count (CBC) change with age in common marmosets (Callithrix jacchus). We assessed blood samples collected during routine physical exams at the Southwest National Primate Research Center and the University of Texas Health San Antonio marmoset colonies from November 2020-November 2021. We found that chemistry and CBC profiles varied based on facility, sex, and age. Significant changes in albumin, phosphorus/creatinine ratio, albumin/globulin ratio, amylase, creatinine, lymphocyte percent, hematocrit, granulocytes percent, lymphocytes, hemoglobin, red cell distribution width, and platelet distribution width were all reported with advancing age. Aged individuals also demonstrated evidence for changes in liver, kidney, and immune system function compared with younger individuals. Our results suggest there may be regular changes associated with healthy aging in marmosets that are outside of the range typically considered as normal values for healthy young individuals, indicating the potential need for redefined healthy ranges for clinical biomarkers in aged animals. Identifying animals that exhibit values outside of this defined healthy aging reference will allow more accurate diagnostics and treatments for aging colonies.

Timing of the luteal-placental shift is delayed with additional fetuses in litter-bearing callitrichid monkeys, Saguinus oedipus and Callithrix jacchus.

The luteal-placental shift is an important milestone of mammalian pregnancy signifying when endocrine control of pregnancy shifts from the corpus luteum of the ovary to the placenta. The corpus luteum is maintained by chorionic gonadotropin (CG). Upon sufficient placental maturation, CG production wanes, the corpus luteum involutes, and control is shifted to the placenta, one consequence of which is a midgestational rise in glucocorticoid production, especially cortisol and cortisone, by both mother and fetus. Glucocorticoids are involved in initiating parturition, prenatal programming of offspring phenotype, and maturing fetal organs. Limited evidence from human pregnancy suggests that the timing of this shift is delayed in twin pregnancies, but little is known about the timing of the luteal-placental shift in litter-bearing monkeys from the primate family Callitrichidae. Here we provide evidence from cotton-top tamarins (Saguinus oedipus) and common marmosets (Callithrix jacchus) of longer duration of elevated CG associated with multiple infant births compared to single births. Urinary profiles from cotton-top tamarins demonstrate that the decline of the extended elevation of CG precedes the onset of the midpregnancy sustained rise in glucocorticoids; this shift occurs later with an increase from one to two fetuses carried to term. In the common marmoset, the onset of the sustained rise of glucocorticoids in maternal urine is also delayed with an increase in infant number. Total urinary glucocorticoid levels during the last half of gestation increase monthly but do not differ by infant number. The significant delay in the luteal-placental shift suggests a longer period of placental maturation is needed to support a greater number of fetuses.

Determination of dexamethasone dose for cortisol suppression in adult common marmosets (Callithrix jacchus).

We conducted a dose-response study of dexamethasone to investigate an optimal dexamethasone suppression test for common marmosets. Twelve marmosets received 0.1, 0.5, or 1.0 mg/kg dexamethasone. Doses of 0.5 and 1.0 mg/kg both suppressed endogenous cortisol for at least 18 h with greater individual variability in the lower 0.5 mg/kg dose.

Feasibility of fecal microbiota transplantation via oral gavage to safely alter gut microbiome composition in marmosets.

Disruption of microbial communities within human hosts has been associated with infection, obesity, cognitive decline, cancer risk and frailty, suggesting that microbiome-targeted therapies may be an option for improving healthspan and lifespan. The objectives of this study were to determine the feasibility of delivering fecal microbiota transplants (FMTs) to marmosets via oral gavage and to evaluate if alteration of the gut microbiome post-FMT could be achieved. This was a prospective study of marmosets housed at the Barshop Institute for Longevity and Aging Studies in San Antonio, Texas. Eligible animals included healthy young adult males (age 2-5 years) with no recent medication use. Stool from two donors was combined and administered in 0.5 ml doses to five young recipients once weekly for 3 weeks. Safety outcomes and alterations in the gut microbiome composition via 16S ribosomal RNA sequencing were compared at baseline and monthly up to 6 months post-FMT. Overall, significant differences in the percent relative abundance was seen in FMT recipients at the phylum and family levels from baseline to 1 month and baseline to 6 months post-FMT. In permutational multivariate analysis of variance analyses, treatment status (donor vs. recipient) (p = .056) and time course (p = .019) predicted ß diversity (p = .056). The FMT recipients did not experience any negative health outcomes over the course of the treatment. FMT via oral gavage was safe to administer to young adult marmosets. The marmoset microbiome may be altered by FMT; however, progressive changes in the microbiome are strongly driven by the host and its baseline microbiome composition.

Maternal weight affects placental DNA methylation of genes involved in metabolic pathways in the common marmoset monkey (Callithrix jacchus).

Accumulating evidence suggests that dysregulation of placental DNA methylation (DNAm) is a mechanism linking maternal weight during pregnancy to metabolic programming outcomes. The common marmoset, Callithrix jaccus, is a platyrrhine primate species that has provided much insight into studies of the primate placenta, maternal condition, and metabolic programming, yet the relationships between maternal weight and placental DNAm are unknown. Here, we report genome-wide DNAm from term marmoset placentas using reduced representation bisulfite sequencing. We identified 74 genes whose DNAm pattern is associated with maternal weight during gestation. These genes are predominantly involved in energy metabolism and homeostasis, including the regulation of glycolytic and lipid metabolic processes pathways.

Noninvasive cardiac psychophysiology as a tool for translational science with marmosets.

The importance of marmosets for comparative and translational science has grown in recent years because of their relatively rapid development, birth cohorts of twins, family social structure, and genetic tractability. Despite this, they remain understudied in investigations of affective processes. In this methodological note, we establish the validity of using noninvasive commercially available equipment to record cardiac physiology and compute indices of autonomic nervous system activity-a major component of affective processes. Specifically, we recorded electrocardiogram and impedance cardiogram, from which we derived heart rate, respiration rate, measures of high-frequency heart rate variability (indices of parasympathetic autonomic nervous system activity), and ventricular contractility (an index of sympathetic autonomic nervous system activity). Our methods produced physiologically plausible data, and further, animals with increased heart rates during testing were also more reactive to isolation from their social partner and presentation of novel objects, though no relationship was observed between reactivity and specific indices of parasympathetic or sympathetic nervous system activity.

Age-related changes in the marmoset gut microbiome.

The gut microbiome is known to play a significant role in human health but its role in aging remains unclear. The objective of this study was to compare the gut microbiome composition between young adult and geriatric non-human primates (marmosets) as a model of human health and disease. Stool samples were collected from geriatric (8+ years) and young adult males (2-5 years). Stool 16S ribosomal RNA V4 sequences were amplified and sequenced on the Illumina MiSeq platform. Sequences were clustered into operational taxonomic units and classified via Mothur's Bayesian classifier referenced against the Greengenes database. A total of 10 young adult and 10 geriatric marmosets were included. Geriatric marmosets had a lower mean Shannon diversity compared with young marmosets (3.15 vs. 3.46; p = 0.0191). Geriatric marmosets had a significantly higher mean abundance of Proteobacteria (0.22 vs. 0.09; p = 0.0233) and lower abundance of Firmicutes (0.15 vs. 0.19; p = 0.0032) compared with young marmosets. Geriatric marmosets had a significantly higher abundance of Succinivibrionaceae (0.16 vs. 0.01; p = 0.0191) and lower abundance of Porphyromonadaceae (0.07 vs. 0.11; p = 0.0494). In summary, geriatric marmosets had significantly altered microbiome diversity and composition compared with young adult marmosets. Further studies are needed to test microbiome-targeted therapies to improve healthspan and lifespan.

Long-term treatment with the mTOR inhibitor rapamycin has minor effect on clinical laboratory markers in middle-aged marmosets.

Interventions to extend lifespan and improve health with increasing age would have significant impact on a growing aged population. There are now several pharmaceutical interventions that extend lifespan in laboratory rodent models with rapamycin, an inhibitor of mechanistic target of rapamycin (mTOR) being the most well studied. In this study, we report on the hematological effects in a cohort of middle-aged common marmosets (Callithrix jacchus) that were enrolled in a study to test the effects of daily rapamycin treatment on aging in this species. In addition, we assessed whether sex was a significant factor in either baseline assessment or as an interaction with rapamycin treatment. Among our cohort at baseline, we found few differences in either basic morphology or hematological markers of blood cell counts, metabolism or inflammation between male and female marmosets. After dosing with rapamycin, surprisingly we found trough blood concentrations of rapamycin were significantly lower in female compared to male marmosets. Despite this pharmacological difference, both sexes had only minor changes in cellular blood counts after 9 months of rapamycin. These data then suggest that the potential clinical hematological side effects of rapamycin are not likely outcomes of long-term rapamycin in relatively healthy, middle-aged marmosets.

Comparative genomics of Bifidobacterium species isolated from marmosets and humans.

The genus Bifidobacterium is purported to have beneficial consequences for human health and is a major component of many gastrointestinal probiotics. Although species of Bifidobacterium are generally at low relative frequency in the adult human gastrointestinal tract, they can constitute high proportions of the gastrointestinal communities of adult marmosets. To identify genes that might be important for the maintenance of Bifidobacterium in adult marmosets, ten strains of Bifidobacterium were isolated from the feces of seven adult marmosets, and their genomes were sequenced. There were six B. reuteri strains, two B. callitrichos strains, one B. myosotis sp. nov. and one B. tissieri sp. nov. among our isolates. Phylogenetic analysis showed that three of the four species we isolated were most closely related to B. bifidum, B. breve and B. longum, which are species found in high abundance in human infants. There were 1357 genes that were shared by at least one strain of B. reuteri, B. callitrichos, B. breve, and B. longum, and 987 genes that were found in all strains of the four species. There were 106 genes found in B. reuteri and B. callitrichos but not in human bifidobacteria, and several of these genes were involved in nutrient uptake. These pathways for nutrient uptake appeared to be specific to Bifidobacterium from New World monkeys. Additionally, the distribution of Bifidobacterium in fecal samples from captive adult marmosets constituted as much as 80% of the gut microbiome, although this was variable between individuals and colonies. We suggest that nutrient transporters may be important for the maintenance of Bifidobacterium during adulthood in marmosets.

Cross-sectional comparison of health-span phenotypes in young versus geriatric marmosets.

The development of the marmoset as a translational model for healthspan and lifespan studies relies on the characterization of health parameters in young and geriatric marmosets. This cross-sectional study examined health phenotypes in marmosets for five domains of interest for human health and aging: mobility, cognition, metabolism, homeostasis, and immune function. Geriatric marmosets were found to have significant executive function impairment when compared to young animals. While geriatric animals did not show gross abnormalities in mobility and measures of locomotion, their types of movement were altered from young animals. Geriatric marmosets had alterations in cardiac function, with significantly increased mean arterial pressures; metabolism, with significantly lower VO2 ; and suppressed immune function. Further, this study sought to characterize and describe histopathology for both young and geriatric healthy marmosets. Overall this study provides a characterization of health parameters for young and geriatric marmosets which will greatly enhance future aging and interventional testing in marmosets.

The common marmoset monkey: avenues for exploring the prenatal, placental, and postnatal mechanisms in developmental programming of pediatric obesity.

Animal models have been critical in building evidence that the prenatal experience and intrauterine environment are capable of exerting profound and permanent effects on metabolic health through developmental programming of obesity. However, despite physiological and evolutionary similarities, nonhuman primate models are relatively rare. The common marmoset monkey ( Callithrix jacchus) is a New World monkey that has been used as a biomedical model for well more than 50 years and has recently been framed as an appropriate model for exploring early-life impacts on later health and disease. The spontaneous, multifactorial, and early-life development of obesity in the common marmoset make it a valuable research model for advancing our knowledge about the role of the prenatal and placental mechanisms involved in developmental programming of obesity. This paper provides a brief overview of obesity in the common marmoset, followed by a discussion of marmoset reproduction and placental characteristics. We then discuss the occurrence and utility of variable intrauterine environments in developmental programming in marmosets. Evidence of developmental programming of obesity will be given, and finally, we put forward future directions and innovations for including the placenta in developmental programming of obesity in the common marmoset.

Serum biomarkers associated with aging and neurodegeneration in common marmosets (Callithrix jacchus).

The common marmoset (Callithrix jacchus), a small South American monkey, is an important nonhuman primate model in the study of aging and age-related neurodegenerative disease, including Alzheimer's disease, Parkinson's disease, and related dementias. Thorough characterization of the wild type marmoset brain agingmodel, including biomarkers of aging and neural degeneration, will further the marmoset's utility in translational research. We measured serum concentration of four key biomarkers of neural degeneration [total tau (T-tau), glial fibrillary acidic protein (GFAP), neurofilament light chain (NfL), and ubiquitin C-terminal hydrolase-L1 (UCH-L1)] via single molecule array from 24 marmosets (female n = 13, male n = 11) ranging in age from 1.3 to 18.7 years. Aged marmosets (>7 years) had significantly higher GFAP, NfL, UCH-L1, and T-tau than adult marmosets. Sex differences were not detected for any of these biomarker concentrations. These data provide an important initial range of reference values for GFAP, NfL, T-tau, and UCH-L1 to evaluate aging and neural health in marmosets, as well as evaluation of therapeutics in clinical models of disease.

Developing the Common Marmoset as a Translational Geroscience Model to Study the Microbiome and Healthy Aging.

Emerging data support associations between the depletion of the healthy gut microbiome and aging-related physiological decline and disease. In humans, fecal microbiota transplantation (FMT) has been used successfully to restore gut microbiome structure and function and to treat C. difficile infections, but its application to healthy aging has been scarcely investigated. The marmoset is an excellent model for evaluating microbiome-mediated changes with age and interventional treatments due to their relatively shorter lifespan and many social, behavioral, and physiological functions that mimic human aging. Prior work indicates that FMT is safe in marmosets and may successfully mediate gut microbiome function and host health. This narrative review (1) provides an overview of the rationale for FMT to support healthy aging using the marmoset as a translational geroscience model, (2) summarizes the prior use of FMT in marmosets, (3) outlines a protocol synthesized from prior literature for studying FMT in aging marmosets, and (4) describes limitations, knowledge gaps, and future research needs in this field.

Protective efficacy of a Zika purified inactivated virus vaccine candidate during pregnancy in marmosets.

Zika virus (ZIKV) infection during pregnancy poses significant threats to maternal and fetal health, leading to intrauterine fetal demise and severe developmental malformations that constitute congenital Zika syndrome (CZS). As such, the development of a safe and effective ZIKV vaccine is a critical public health priority. However, the safety and efficacy of such a vaccine during pregnancy remain uncertain. Historically, the conduct of clinical trials in pregnant women has been challenging. Therefore, clinically relevant animal pregnancy models are in high demand for testing vaccine efficacy. We previously reported that a marmoset pregnancy model of ZIKV infection consistently demonstrated vertical transmission from mother to fetus during pregnancy. Using this marmoset model, we also showed that vertical transmission could be prevented by pre-pregnancy vaccination with Zika purified inactivated virus (ZPIV) vaccine. Here, we further examined the efficacy of ZPIV vaccination during pregnancy. Vaccination during pregnancy elicited virus neutralizing antibody responses that were comparable to those elicited by pre-pregnancy vaccination. Vaccination also reduced placental pathology, viral burden and vertical transmission of ZIKV during pregnancy, without causing adverse effects. These results provide key insights into the safety and efficacy of ZPIV vaccination during pregnancy and demonstrate positive effects of vaccination on the reduction of ZIKV infection, an important advance in preparedness for future ZIKV outbreaks.

Body mass growth in common marmosets: toward a model of pediatric obesity.

While much is known about adult obesity in nonhuman primates, very little is known regarding development of childhood adiposity. As small monkeys that are easy to handle and have a relatively fast life history, common marmoset monkeys (Callithrix jacchus) offer interesting opportunities to examine the question of fat versus lean mass growth in a nonhuman primate. This article provides an overview of our understanding of early life growth in mass in marmoset monkeys, based primarily upon our past 20 years of research, culminating in our recent findings on early life obesity in this species. Common marmosets display variance in early life growth patterns that is related to both pre- and postnatal factors and the marmoset uterine environment is exquisitely designed to reflect resources available for the gestation of multiple offspring, making them an interesting model of developmental programming. We have demonstrated that obesity can be generated in very early life in captive marmosets, with excess adiposity evident by one month of age, making this species a potentially valuable model in which to study pediatric obesity and its sequelae. Birth weight is associated with adiposity in animals vulnerable to obesity. Early life exposure to higher fat diets enhances the chances of postweaning obesity development. However, overall higher food consumption is also associated with obesity development at later ages. One unexpected finding in our studies has been the relatively high body fat percentage of neonatal (12-18%) marmosets suggesting that hypotheses regarding the uniqueness of high human neonatal adiposity merit further examination.

Evaluation of Anesthetic and Cardiorespiratory Effects after Intramuscular Administration of Three Different Doses of Telazol® in Common Marmosets (Callithrix jacchus).

Marmosets' small body size makes anesthesia challenging. Ideally, small volumes of drugs should be administered intramuscularly (i.m.). In addition, dose-dependent sedation and anesthesia are desirable properties for sedatives and anesthetics in marmosets. Telazol® (tiletamine and zolazepam) is highly concentrated, allowing the use of small injection volumes and dose-dependent sedation and anesthesia. A randomized, blinded study with crossover design in ten healthy adult common marmosets (Callithrix jacchus) was performed to evaluate the anesthetic and cardiorespiratory effects of three doses of i.m. Telazol® (respectively, 5, 10, and 15 mg/kg). Depth of anesthesia, cardiorespiratory effects, and induction, immobilization, and recovery times were determined. A significant difference was observed in immobilization time between 5 and 15 mg/kg of Telazol®. In addition, 15 mg/kg of Telazol® resulted in increased recovery times compared to 5 mg/kg. The cardiorespiratory effects during the first 45 min of immobilization were within clinically acceptable limits. The pedal withdrawal reflex was the best indicator of the anesthetic depth.

Efficient generation of marmoset primordial germ cell-like cells using induced pluripotent stem cells.

Reconstitution of germ cell fate from pluripotent stem cells provides an opportunity to understand the molecular underpinnings of germ cell development. Here, we established robust methods for induced pluripotent stem cell (iPSC) culture in the common marmoset (Callithrix jacchus [cj]), allowing stable propagation in an undifferentiated state. Notably, iPSCs cultured on a feeder layer in the presence of a WNT signaling inhibitor upregulated genes related to ubiquitin-dependent protein catabolic processes and enter a permissive state that enables differentiation into primordial germ cell-like cells (PGCLCs) bearing immunophenotypic and transcriptomic similarities to pre-migratory cjPGCs in vivo. Induction of cjPGCLCs is accompanied by transient upregulation of mesodermal genes, culminating in the establishment of a primate-specific germline transcriptional network. Moreover, cjPGCLCs can be expanded in monolayer while retaining the germline state. Upon co-culture with mouse testicular somatic cells, these cells acquire an early prospermatogonia-like phenotype. Our findings provide a framework for understanding and reconstituting marmoset germ cell development in vitro, thus providing a comparative tool and foundation for a preclinical modeling of human in vitro gametogenesis.

Whole Genome Analysis of SNV and Indel Polymorphism in Common Marmosets (Callithrix jacchus).

The common marmoset (Callithrix jacchus) is one of the most widely used nonhuman primate models of human disease. Owing to limitations in sequencing technology, early genome assemblies of this species using short-read sequencing suffered from gaps. In addition, the genetic diversity of the species has not yet been adequately explored. Using long-read genome sequencing and expert annotation, we generated a high-quality genome resource creating a 2.898 Gb marmoset genome in which most of the euchromatin portion is assembled contiguously (contig N50 = 25.23 Mbp, scaffold N50 = 98.2 Mbp). We then performed whole genome sequencing on 84 marmosets sampling the genetic diversity from several marmoset research centers. We identified a total of 19.1 million single nucleotide variants (SNVs), of which 11.9 million can be reliably mapped to orthologous locations in the human genome. We also observed 2.8 million small insertion/deletion variants. This dataset includes an average of 5.4 million SNVs per marmoset individual and a total of 74,088 missense variants in protein-coding genes. Of the 4956 variants orthologous to human ClinVar SNVs (present in the same annotated gene and with the same functional consequence in marmoset and human), 27 have a clinical significance of pathogenic and/or likely pathogenic. This important marmoset genomic resource will help guide genetic analyses of natural variation, the discovery of spontaneous functional variation relevant to human disease models, and the development of genetically engineered marmoset disease models.

Establishing meal patterns by lickometry in the marmoset monkey (Callithrix jacchus): translational applications from the bench to the field and the clinic.

The ability to measure and interpret variables associated with feeding behavior and food intake is essential to a variety of nonhuman primate study modalities. The development of a technique to accurately and efficiently measure food intake and meal patterning in captivity will enhance both the interpretation of foraging behavior in the wild as well as our ability to model clinically relevant human feeding pathologies. In this study, we successfully developed the use of a rodent lickometer system to monitor meal patterning in captive common marmosets. We describe the modifications necessary for this type of instrumentation to be used successfully with marmosets. We define variables of interest that relate to both previous rodent literature and human clinical measures. Finally, we relate our findings to potential translational value for both primate field research and biomedical applications.

The development of obesity begins at an early age in captive common marmosets (Callithrix jacchus).

Animal models to study the causes and consequences of obesity during infancy in humans would be valuable. In this study, we examine the patterns of fat mass gain from birth to 12 months in common marmosets (Callithrix jacchus). Lean and fat mass was measured by quantitative magnetic resonance at 1, 2, 6, and 12 months for 31 marmosets, 15 considered Normal and 16 considered Fat (> 14% body fat) at 12 months. Animals were fed either the regular colony diet mix or a high-fat variation. Subjects classified as Fat at 12 months already had greater lean mass (198.4 +/- 6.2 g vs. 174.0 +/- 6.8 g, P = 0.013) and fat mass (45.5 +/- 5.0 g vs. 24.9 +/- 3.4 g, P = .002) by 6 months. Body mass did not differ between groups prior to 6 months, however, by 1 month, Fat infants had greater percent body fat. Percent body fat decreased between 1 and 12 months in Normal subjects; in Fat subjects, it increased. The high-fat diet was associated with body fat > 14% at 6 months (P = 0.049), but not at 12 months. This shift was due to three subjects on the normal diet changing from Normal to Fat between 6 and 12 months. Although maternal prepregnancy adiposity did not differ, overall, between Normal and Fat subjects, the subjects Normal at 6 and Fat at 12 months all had Fat mothers. Therefore, diet and maternal obesity appear to have potentially independent effects that may also vary with developmental age. Although birth weight did not differ between groups, it was associated with fat mass gain from 1 to 6 months in animals with > 14% body fat at 6 months of age (r = 0.612, P = 0.026); but not in 6-month-old animals with < 14% body fat (r = -0.012, P = 0.964). Excess adiposity in captive marmosets develops by 1 month. Birth weight is associated with adiposity in animals vulnerable to obesity.