Assessing cerebrovascular reactivity (CVR) in rhesus macaques (Macaca mulatta) using a hypercapnic challenge and pseudo-continuous arterial spin labeling (pCASL).

Cerebrovascular reactivity (CVR) is a measure of cerebral small vessels' ability to respond to changes in metabolic demand and can be quantified using magnetic resonance imaging (MRI) coupled with a vasoactive stimulus. Reduced CVR occurs with neurodegeneration and is associated with cognitive decline. While commonly measured in humans, few studies have evaluated CVR in animal models. Herein, we describe methods to induce hypercapnia in rhesus macaques (Macaca mulatta) under gas anesthesia to measure cerebral blood flow (CBF) and CVR using pseudo-continuous arterial spin labeling (pCASL). Fifteen (13 M, 2 F) adult rhesus macaques underwent pCASL imaging that included a baseline segment (100% O2) followed by a hypercapnic challenge (isoflurane anesthesia with 5% CO2, 95% O2 mixed gas). Relative hypercapnia was defined as an end-tidal CO2 (ETCO2) ≥5 mmHg above baseline ETCO2. The mean ETCO2 during the baseline segment of the pCASL sequence was 34 mmHg (range: 23-48 mmHg). During this segment, mean whole-brain CBF was 51.48 ml/100g/min (range: 21.47-77.23 ml/100g/min). Significant increases (p<0.0001) in ETCO2 were seen upon inspiration of the mixed gas (5% CO2, 95% O2). The mean increase in ETCO2 was 8.5 mmHg and corresponded with a mean increase in CBF of 37.1% (p<0.0001). The mean CVR measured was 4.3%/mmHg. No anesthetic complications occurred as a result of the CO2 challenge. Our methods were effective at inducing a state of relative hypercapnia that corresponds with a detectable increase in whole brain CBF using pCASL MRI. Using these methods, a CO2 challenge can be performed in conjunction with pCASL imaging to evaluate CBF and CVR in rhesus macaques. The measured CVR in rhesus macaques is comparable to human CVR highlighting the translational utility of rhesus macaques in neuroscience research. These methods present a feasible means to measure CVR in comparative models of neurodegeneration and cerebrovascular dysfunction.

Total-Body Irradiation Alters White Matter Volume and Microstructural Integrity in Rhesus Macaques.

PURPOSE: Long-term survivors of brain irradiation can experience irreversible injury and cognitive impairment. T1-weighted and diffusion tensor magnetic resonance imaging (MRI) are used to evaluate brain volume and white matter (WM) microstructure in neurodevelopmental and neurodegenerative conditions. The goal of this study was to evaluate the long-term effects of single-dose total-body irradiation (TBI) or TBI with 5% partial-body sparing on brain volumetrics and WM integrity in macaques. METHODS AND MATERIALS: We used MRI scans from a cohort of male rhesus macaques (age range, 3.6-22.8 years) to compare global and regional brain volumes and WM diffusion in survivors of TBI (T1-weighted, n = 137; diffusion tensor imaging, n = 121; dose range, 3.5-10 Gy) with unirradiated controls (T1-weighted, n = 48; diffusion tensor imaging, n = 38). RESULTS: In all regions of interest, radiation affected age-related changes in fractional anisotropy, which tended to increase across age in both groups but to a lesser extent in the irradiated group (interaction P < .01). Depending on the region of interest, mean diffusivity decreased or remained the same across age in unirradiated animals, whereas it increased or did not change in irradiated animals. The increases in mean diffusivity were driven by changes in radial diffusivity, which followed similar trends across age. Axial diffusivity did not differ by irradiation status. Age-related changes in relative volumes in controls reflected normal trends in humans, with increasing WM and decreasing gray matter until middle age. Cerebrospinal fluid (CSF) volume did not differ across age in controls. WM volume was lower and CSF volume was higher in young irradiated macaques. WM volume was similar between groups, and CSF volume lower in older irradiated macaques. Gray matter volume was unaffected by radiation. CONCLUSIONS: TBI results in delayed WM expansion and long-term disruption of WM integrity. Diffusion changes suggest that myelin injury in WM is a hallmark of late-delayed radiation-induced brain injury.

Dose-Dependent Testicular Injury and Recovery after Total-Body Irradiation in Rhesus Monkeys.

Testicular injury is a well-documented acute effect of radiation exposure, though little is known about recovery years after irradiation, especially at higher doses. We examined the testes from 143 irradiated and control male rhesus monkeys, who were part of the Radiation Late Effects Cohort over a four-year period. Irradiated animals were exposed to doses ranging from 3.5 to 8.5 Gy of total-body irradiation. The testes were assessed using computed tomography (CT) volumetry, serum testosterone, and histology for deceased members of the cohort. Irradiated animals exhibited dose-dependent testicular atrophy as well as decreased serum testosterone during the winter breeding season when compared to age-matched unirradiated controls. No significant difference in summer testosterone levels was observed. Volumetric and histologic evidence of testicular recovery was present approximately three years postirradiation for animals who received ≤8 Gy. The study demonstrates dose-dependent testicular injury after total-body irradiation and provides evidence for volumetric and spermatogonial recovery even at lethal doses of total-body irradiation in rhesus monkeys.

Long-Term Immunological Consequences of Radiation Exposure in a Diverse Cohort of Rhesus Macaques.

PURPOSE: The aim of this study was to develop an improved understanding of the delayed immunologic effects of acute total body irradiation (TBI) using a diverse cohort of nonhuman primates as a model for an irradiated human population. METHODS AND MATERIALS: Immune recovery was evaluated in 221 rhesus macaques either left unirradiated (n = 36) or previously irradiated (n = 185) at 1.1 to 8.5 Gy TBI (median, 6.5 Gy) when aged 2.1 to 15.5 years (median, 4.2 years). Blood was drawn annually for up to 5 years total between 0.5 and 14.3 years after exposure. Blood was analyzed by complete blood count, immunophenotyping of monocytes, dendritic cells (DC) and lymphocytes by flow cytometry, and signal joint T-cell receptor exclusion circle quantification in isolated peripheral blood CD4 and CD8 T cells. Animals were categorized by age, irradiation status, and time since irradiation. Sex-adjusted means of immune metrics were evaluated by generalized estimating equation models to identify cell populations altered by TBI. RESULTS: Overall, the differences between irradiated and nonirradiated animals were subtle and largely restricted to younger animals and select cell populations. Subsets of monocytes, DC, T cells, and B cells showed significant interaction effects between radiation dose and age after adjustment for sex. Irradiation at a young age caused transient increases in the percentage of peripheral blood myeloid DC and dose-dependent changes in monocyte balance for at least 5 years after TBI. TBI also led to a sustained decrease in the percentage of circulating memory B cells. Young irradiated animals exhibited statistically significant and prolonged disruption of the naïve/effector memory/central memory CD4 and CD8 T-cell equilibrium and exhibited a dose-dependent increase in thymopoiesis for 2 to 3 years after exposure. CONCLUSIONS: This study indicates TBI subtly but significantly alters the circulating proportions of cellular mediators of adaptive immune memory for several years after irradiation, especially in macaques under 5 years of age and those receiving a high dose of radiation.

[18F]KS1, a novel ascorbate-based ligand images ROS in tumor models of rodents and nonhuman primates.

Over production of reactive oxygen species (ROS) caused by altered redox regulation of signaling pathways is common in many types of cancers. While PET imaging is recognized as the standard tool for cancer imaging, there are no clinically-approved PET radiotracers for ROS-imaging in cancer diagnosis and treatment. An ascorbate-based radio ligand promises to meet this urgent need. Our laboratory recently synthesized [18F] KS1, a fluoroethoxy furanose ring-containing ascorbate derivative, to track ROS in prostate tumor-bearing mice. Here we report cell uptake assays of [18F]KS1 with different ROS-regulating agents, PET imaging in head and neck squamous cell carcinoma (HNSCC) mice, and doxorubicin-induced rats; PET imaging in healthy and irradiated hepatic tumor-bearing rhesus to demonstrate its translational potential. Our preliminary evaluations demonstrated that KS1 do not generate ROS in tumor cells at tracer-level concentrations and tumor-killing properties at pharmacologic doses. [18F]KS1 uptake was low in HNSCC pretreated with ROS blockers, and high with ROS inducers. Tumors in high ROS-expressing SCC-61 took up significantly more [18F]KS1 than rSCC-61 (low-ROS expressing HNSCC); high uptake in doxorubicin-treated rats compared to saline-treated controls. Rodent biodistribution and PET imaging of [18F]KS1 in healthy rhesus monkeys demonstrated its favorable safety, pharmacokinetic properties with excellent washout profile, within 3.0 h of radiotracer administration. High uptake of [18F]KS1 in liver tumor tissues of the irradiated hepatic tumor-bearing monkey showed target selectivity. Our strong data in vitro, in vivo, and ex vivo here supports the high translational utility of [18F]KS1 to image ROS.

Total-Body Irradiation Is Associated With Increased Incidence of Mesenchymal Neoplasia in a Radiation Late Effects Cohort of Rhesus Macaques (Macaca mulatta).

PURPOSE: Cancer is a severe delayed effect of acute radiation exposure. Total-body irradiation has been associated with an increased risk of solid cancer and leukemia in Japanese atomic bomb survivors, and secondary malignancies, such as sarcoma, are a serious consequence of cancer radiation therapy. The radiation late effects cohort (RLEC) of rhesus macaques (Macaca mulatta) is a unique resource of more than 200 animals for studying the long-term consequences of total-body irradiation in an animal model that closely resembles humans at the genetic and physiologic levels. METHODS AND MATERIALS: Using clinical records, clinical imaging, histopathology, and immunohistochemistry, this retrospective study characterized the incidence of neoplasia in the RLEC. RESULTS: Since 2007, 61 neoplasms in 44 of 239 irradiated animals were documented (18.4% of the irradiated population). Only 1 neoplasm was diagnosed among the 51 nonirradiated controls of the RLEC (2.0%). The most common malignancies in the RLEC were sarcomas (38.3% of diagnoses), which are rare neoplasms in nonirradiated macaques. The most common sarcomas included malignant nerve sheath tumors and malignant glomus tumors. Carcinomas were less common (19.7% of diagnoses), and consisted primarily of renal cell and hepatocellular carcinomas. Neoplasia occurred in most major body systems, with the skin and subcutis being the most common site (40%). RNA analysis showed similarities in transcriptional profiles between RLEC and human malignant nerve sheath tumors. CONCLUSIONS: This study indicates that total-body irradiation is associated with an increased incidence of neoplasia years following irradiation, at more than double the incidence described in aging, nonirradiated animals, and promotes tumor histotypes that are rarely observed in nonirradiated, aging rhesus macaques.

Hypertension promotes microbial translocation and dysbiotic shifts in the fecal microbiome of nonhuman primates.

Accumulating evidence indicates a link between gut barrier dysfunction and hypertension. However, it is unclear whether hypertension causes gut barrier dysfunction or vice versa and whether the gut microbiome plays a role. To understand this relationship, we first cross-sectionally examined 153 nonhuman primates [NHPs; Chlorocebus aethiops sabaeus; mean age, 16 ± 0.4 yr; 129 (84.3%) females] for cardiometabolic risk factors and gut barrier function biomarkers. This analysis identified blood pressure and age as specific factors that independently associated with microbial translocation. We then longitudinally tracked male, age-matched spontaneously hypertensive NHPs (Macaca mulatta) to normotensives (n = 16), mean age of 5.8 ± 0.5 yr, to confirm hypertension-related gut barrier dysfunction and to explore the role of microbiome by comparing groups at baseline, 12, and 27 mo. Collectively, hypertensive animals in both studies showed evidence of gut barrier dysfunction (i.e., microbial translocation), as indicated by higher plasma levels of lipopolysaccharide-binding protein (LBP)-1, when compared with normotensive animals. Furthermore, plasma LBP-1 levels were correlated with diastolic blood pressure, independent of age and other health markers, suggesting specificity of the effect of hypertension on microbial translocation. In over 2 yr of longitudinal assessment, hypertensive animals had escalating plasma levels of LBP-1 and greater bacterial gene expression in mesenteric lymph nodes compared with normotensive animals, confirming microbes translocated across the intestinal barrier. Concomitantly, we identified distinct shifts in the gut microbial signature of hypertensive versus normotensive animals at 12 and 27 mo. These results suggest that hypertension contributes to microbial translocation in the gut and eventually unhealthy shifts in the gut microbiome, possibly contributing to poor health outcomes, providing further impetus for the management of hypertension.NEW & NOTEWORTHY Hypertension specifically had detrimental effects on microbial translocation when age and metabolic syndrome criteria were evaluated as drivers of cardiovascular disease in a relevant nonhuman primate model. Intestinal barrier function exponentially decayed over time with chronic hypertension, and microbial translocation was confirmed by detection of more microbial genes in regional draining lymph nodes. Chronic hypertension resulted in fecal microbial dysbiosis and elevations of the biomarker NT-proBNP. This study provides insights on the barrier dysfunction, dysbiosis, and hypertension in controlled studies of nonhuman primates. Our study includes a longitudinal component comparing naturally occurring hypertensive to normotensive primates to confirm microbial translocation and dysbiotic microbiome development. Hypertension is an underappreciated driver of subclinical endotoxemia that can drive chronic inflammatory diseases.

Long-Term Recovery of the Adaptive Immune System in Rhesus Macaques After Total Body Irradiation.

PURPOSE: Ionizing radiation causes acute damage to hematopoietic and immune cells, but the long-term immunologic consequences of irradiation are poorly understood. We therefore performed a prospective study of the delayed immune effects of radiation using a rhesus macaque model. METHODS AND MATERIALS: Ten macaques received 4 Gy high-energy x-ray total body irradiation (TBI) and 6 control animals received sham irradiation. TBI caused transient lymphopenia that resolved over several weeks. Once white blood cell counts recovered, flow cytometry was used to immunophenotype the circulating adaptive immune cell populations 4, 9, and 21 months after TBI. Data were fit using a mixed-effects model to determine age-dependent, radiation-dependent, and interacting effects. T cell receptor (TCR) sequencing and quantification of TCR Excision Circles were used to determine relative contributions of thymopoiesis and peripheral expansion to T cell repopulation. Two years after TBI, the cohort was vaccinated with a 23-valent pneumococcal polysaccharide vaccine and a tetravalent influenza hemagglutinin vaccine. RESULTS: Aging, but not TBI, led to significant changes in the frequencies of dendritic cells, CD4 and CD8 T cells, and B cells. However, irradiated animals exhibited increased frequencies of central memory T cells and decreased frequencies of naïve T cells. These consequences of irradiation were time-dependent and more prolonged in the CD8 T cell population. Irradiation led to transient increases in CD8+ T cell TCR Excision Circles and had no significant effect on TCR sequence entropy, indicating T cell recovery was partially mediated by thymopoiesis. Animals that were irradiated and then vaccinated showed normal immunoglobulin G binding and influenza neutralization titers in response to the 4 protein antigens but weaker immunoglobulin G binding titers to 10 of the 23 polysaccharide antigens. CONCLUSIONS: These findings indicate that TBI causes subtle but long-lasting immune defects that are evident years after recovery from lymphopenia.

Detection and quantification of renal fibrosis by computerized tomography.

OBJECTIVES: Reliable biomarkers for renal fibrosis are needed for clinical care and for research. Existing non-invasive biomarkers are imprecise, which has limited their utility. METHODS: We developed a method to quantify fibrosis by subject size-adjusted CT Hounsfield units. This was accomplished using CT measurements of renal cortex in previously irradiated non-human primates. RESULTS: Renal cortex mean CT Hounsfield units that were adjusted for body size had a very good direct correlation with renal parenchymal fibrosis, with an area under the curve of 0.93. CONCLUSIONS: This metric is a promising and simple non-invasive biomarker for renal fibrosis.

Late effects of total body irradiation on hematopoietic recovery and immune function in rhesus macaques.

While exposure to radiation can be lifesaving in certain settings, it can also potentially result in long-lasting adverse effects, particularly to hematopoietic and immune cells. This study investigated hematopoietic recovery and immune function in rhesus macaques Cross-sectionally (at a single time point) 2 to 5 years after exposure to a single large dose (6.5 to 8.4 Gray) of total body radiation (TBI) derived from linear accelerator-derived photons (2 MeV, 80 cGy/minute) or Cobalt 60-derived gamma irradiation (60 cGy/min). Hematopoietic recovery was assessed through measurement of complete blood counts, lymphocyte subpopulation analysis, and thymus function assessment. Capacity to mount specific antibody responses against rabies, Streptococcus pneumoniae, and tetanus antigens was determined 2 years after TBI. Irradiated macaques showed increased white blood cells, decreased platelets, and decreased frequencies of peripheral blood T cells. Effects of prior radiation on production and export of new T cells by the thymus was dependent on age at the time of analysis, with evidence of interaction with radiation dose for CD8+ T cells. Irradiated and control animals mounted similar mean antibody responses to proteins from tetanus and rabies and to 10 of 11 serotype-specific pneumococcal polysaccharides. However, irradiated animals uniformly failed to make antibodies against polysaccharides from serotype 5 pneumococci, in contrast to the robust responses of non-irradiated controls. Trends toward decreased serum levels of anti-tetanus IgM and slower peak antibody responses to rabies were also observed. Taken together, these data show that dose-related changes in peripheral blood cells and immune responses to both novel and recall antigens can be detected 2 to 5 years after exposure to whole body radiation. Longer term follow-up data on this cohort and independent validation will be helpful to determine whether these changes persist or whether additional changes become evident with increasing time since radiation, particularly as animals begin to develop aging-related changes in immune function.

Cortical Thinning and Structural Bone Changes in Non-Human Primates after Single-Fraction Whole-Chest Irradiation.

Stereotactic body radiation therapy (SBRT) is associated with an increased risk of vertebral compression fracture. While bone is typically considered radiation resistant, fractures frequently occur within the first year of SBRT. The goal of this work was to determine if rapid deterioration of bone occurs in vertebrae after irradiation. Sixteen male rhesus macaque non-human primates (NHPs) were analyzed after whole-chest irradiation to a midplane dose of 10 Gy. Ages at the time of exposure varied from 45-134 months. Computed tomography (CT) scans were taken 2 months prior to irradiation and 2, 4, 6 and 8 months postirradiation for all animals. Bone mineral density (BMD) and cortical thickness were calculated longitudinally for thoracic (T) 9, lumbar (L) 2 and L4 vertebral bodies; gross morphology and histopathology were assessed per vertebra. Greater mortality (related to pulmonary toxicity) was noted in NHPs <50 months at time of exposure versus NHPs >50 months ( P = 0.03). Animals older than 50 months at time of exposure lost cortical thickness in T9 by 2 months postirradiation ( P = 0.0009), which persisted to 8 months. In contrast, no loss of cortical thickness was observed in vertebrae out-of-field (L2 and L4). Loss of BMD was observed by 4 months postirradiation for T9, and 6 months postirradiation for L2 and L4 ( P < 0.01). For NHPs younger than 50 months at time of exposure, both cortical thickness and BMD decreased in T9, L2 and L4 by 2 months postirradiation ( P < 0.05). Regions that exhibited the greatest degree of cortical thinning as determined from CT scans also exhibited increased porosity histologically. Rapid loss of cortical thickness was observed after high-dose chest irradiation in NHPs. Younger age at time of exposure was associated with increased pneumonitis-related mortality, as well as greater loss of both BMD and cortical thickness at both in- and out-of-field vertebrae. Older NHPs exhibited rapid loss of BMD and cortical thickness from in-field vertebrae, but only loss of BMD in out-of-field vertebrae. Bone is sensitive to high-dose radiation, and rapid loss of bone structure and density increases the risk of fractures.

Post-Irradiation Treatment with a Superoxide Dismutase Mimic, MnTnHex-2-PyP5+, Mitigates Radiation Injury in the Lungs of Non-Human Primates after Whole-Thorax Exposure to Ionizing Radiation.

Radiation injury to the lung is the result of acute and chronic free radical formation, and there are currently few effective means of mitigating such injury. Studies in rodents indicate that superoxide dismutase mimetics may be effective in this regard; however, studies in humans or large animals are lacking. We hypothesized that post-exposure treatment with the lipophilic mitochondrial superoxide dismutase mimetic, MnTnHex-2-PyP5+ (hexyl), would reduce radiation-induced pneumonitis and fibrosis in the lungs of nonhuman primates. Rhesus monkeys (Macaca mulatta) received 10 Gy whole thorax irradiation, 10 Gy + hexyl treatment, sham irradiation, or sham irradiation + hexyl. Hexyl was given twice daily, subcutaneously, at 0.05 mg/kg, for 2 months. Animals were monitored daily, and respiratory rates, pulse oximetry, hematology and serum chemistry panels were performed weekly. Computed tomography scans were performed at 0, 2, and 4 months after irradiation. Supportive fluid therapy, corticosteroids, analgesics, and antibiotics were given as needed. All animals were humanely euthanized 4.5 months after irradiation, and pathologic assessments were made. Multifocal, progressive lung lesions were seen at 2 and 4 months in both irradiated groups. Hexyl treatment delayed the onset of radiation-induced lung lesions, reduced elevations of respiratory rate, and reduced pathologic increases in lung weight. No adverse effects of hexyl treatment were found. These results demonstrate (1) development of a nonhuman primate model of radiation-induced lung injury, (2) a significant mitigating effect of hexyl treatment on lung pathology in this model, and (3) no evidence for toxicity of hexyl at the dose studied.

Long-Term Cognitive Functioning in Single-Dose Total-Body Gamma-Irradiated Rhesus Monkeys ( Macaca mulatta ).

In this study, the effects of a potentially lethal radiation exposure on the brain for long-term cognitive sequelae were investigated using Rhesus macaques ( Macaca mulatta ) adopted from other facilities after analysis of acute radiation response via the Centers for Medical Countermeasures against Radiation (CMCR) network. Fifty-nine animals were given the opportunity to participate in cognitive cage-side testing. The animals that received single-dose gamma irradiation were significantly less likely to engage in cognitive testing than the controls, suggesting that irradiated animals may have differences in cognitive ability. Five irradiated (6.75-8.05 Gy) and three naïve control animals self-selected, were extensively trained and administered a simple visual discrimination with reversal (SVD+R) task 2-3 times per week for 11-18 months. Each session consisted of 30 trials in which the animals were required to choose the correct visual stimulus for a food reward. After the initial presentation, the stimulus that signaled the presence of food was twice reversed once the animal reached criterion (90% accuracy across four consecutive sessions). While the limited sample size precluded definitive statistical analysis, irradiated animals took longer to reach the criterion subsequent to reversal than did control animals, suggesting a relative deficiency in cognitive flexibility. These results provide preliminary data supporting the potential use of a nonhuman primate model to study radiation-induced, late-delayed cognitive deficits.

Late Effects of Total-Body Gamma Irradiation on Cardiac Structure and Function in Male Rhesus Macaques.

Heart disease is an increasingly recognized, serious late effect of radiation exposure, most notably among breast cancer and Hodgkin's disease survivors, as well as the Hiroshima and Nagasaki atomic bomb survivors. The purpose of this study was to evaluate the late effects of total-body irradiation (TBI) on cardiac morphology, function and selected circulating biomarkers in a well-established nonhuman primate model. For this study we used male rhesus macaques that were exposed to a single total-body dose of ionizing gamma radiation (6.5-8.4 Gy) 5.6-9.7 years earlier at ages ranging from ∼3-10 years old and a cohort of nonirradiated controls. Transthoracic echocardiography was performed annually for 3 years on 20 irradiated and 11 control animals. Myocardium was examined grossly and histologically, and myocardial fibrosis/collagen was assessed microscopically and by morphometric analysis of Masson's trichrome-stained sections. Serum/plasma from 27 irradiated and 13 control animals was evaluated for circulating biomarkers of cardiac damage [N-terminal pro B-type natriuretic protein (nt-proBNP) and troponin-I], inflammation (CRP, IL-6, MCP-1, sICAM) and microbial translocation [LPS-binding protein (LBP) and sCD14]. A higher prevalence of histological myocardial fibrosis was observed in the hearts obtained from the irradiated animals (9/14) relative to controls (0/3) (P = 0.04, χ(2)). Echocardiographically determined left ventricular end diastolic and systolic diameters were significantly smaller in irradiated animals (repeated measures ANOVA, P < 0.001 and P < 0.008, respectively). Histomorphometric analysis of trichrome-stained sections of heart tissue demonstrated ∼14.9 ± 1.4% (mean ± SEM) of myocardial area staining for collagen in irradiated animals compared to 9.1 ± 0.9 % in control animals. Circulating levels of MCP-1 and LBP were significantly higher in irradiated animals (P < 0.05). A high incidence of diabetes in the irradiated animals was associated with higher plasma triglyceride and lower HDLc but did not appear to be associated with cardiovascular phenotypes. These results demonstrate that single total-body doses of 6.5-8.4 Gy produced long-term effects including a high incidence of myocardial fibrosis, reduced left ventricular diameter and elevated systemic inflammation. Additional prospective studies are required to define the time course and mechanisms underlying radiation-induced heart disease in this model.

Type 2 Diabetes is a Delayed Late Effect of Whole-Body Irradiation in Nonhuman Primates.

One newly recognized consequence of radiation exposure may be the delayed development of diabetes and metabolic disease. We document the development of type 2 diabetes in a unique nonhuman primate cohort of monkeys that were whole-body irradiated with high doses (6.5-8.4 Gy) 5-9 years earlier. We report here a higher prevalence of type 2 diabetes in irradiated monkeys compared to age-matched nonirradiated monkeys. These irradiated diabetic primates demonstrate insulin resistance and hypertriglyceridemia, however, they lack the typical obese presentation of primate midlife diabetogenesis. Surprisingly, body composition analyses by computed tomography indicated that prior irradiation led to a specific loss of visceral fat mass. Prior irradiation led to reductions in insulin signaling effectiveness in skeletal muscle and higher monocyte chemoattractant protein 1 levels, indicative of increased inflammation. However, there was an absence of large defects in pancreatic function with radiation exposure, which has been documented previously in animal and human studies. Monkeys that remained healthy and did not become diabetic in the years after irradiation were significantly leaner and smaller, and were generally smaller and younger at the time of exposure. Irradiation also resulted in smaller stature in both diabetic and nondiabetic monkeys, compared to nonirradiated age-matched controls. Our study demonstrates that diabetogenesis postirradiation is not a consequence of disrupted adipose accumulation (generalized or in ectopic depots), nor generalized pancreatic failure, but suggests that peripheral tissues such as the musculature are impaired in their response to insulin exposure. Ongoing inflammation in these animals appears to be a consequence of radiation exposure and can interfere with insulin signaling. The reasons that some animals remain protected from diabetes as a late effect of irradiation are not clear, but may be related to body size. The translational relevance for these results suggest that muscle may be an important and underappreciated target organ for the delayed late effect of whole-body irradiation, leading to increased risk of insulin resistance and diabetes development.

Assessment of Multiplate platelet aggregometry using citrate, heparin or hirudin in Rhesus macaques.

Electrical impedance aggregometry (EIA) has gained popularity in clinical and research applications. Nonhuman primates are used to study disease and drug-related mechanisms that affect hemostasis, therefore establishing normal EIA parameters are necessary. The anticoagulants sodium heparin, hirudin and sodium citrate and three agonists, ADP, ASPI, and collagen were evaluated. Whole blood from 12 adult male rhesus macaques was collected to evaluate anticoagulants, sodium heparin, hirudin and sodium citrate using three agonists (ADP, ASPI and collagen), on the Multiplate® 5.0 Analyzer. Platelet function was reported for three parameters: Area under the curve (AUC), aggregation, and aggregation velocity. There was a significant difference in mean AUC between citrate and heparin samples, and citrate and hirudin samples regardless of the agonist used. There was no difference in AUC between heparin and hirudin. ADP-activated samples showed an increase in impedance with hirudin samples compared to citrate. Furthermore, heparin and hirudin out-perform citrate as the anticoagulant for EIA in the macaque. Finally, this study demonstrates the utility of the Multiplate® system in this model and provides important insight into anticoagulant choice when using EIA.

Monkey see, monkey do: contagious itch in nonhuman primates.

"Contagious itch" has been anecdotally reported and recently confirmed in a controlled setting in humans. Here, we investigated in adult rhesus macaques whether 'contagious itch' occurs spontaneously in monkeys. In a first experiment, the latency to scratch following cage-mate scratching was observed in pair-housed adult rhesus macaques. Scratching increased within the first 60 s and subsequently declined. In a second experiment, scratching behavior was recorded for individually caged adult rhesus macaques which where shown videos of monkeys scratching, but also neutral stimuli. A greater frequency of scratching was observed when monkeys viewed a video sequence of another monkey scratching as well as during the neutral stimulus immediately following the monkey scratching segment. In conclusion, viewing other monkeys scratching significantly increased scratching behavior in adult rhesus macaques.

Growth hormone mitigates against lethal irradiation and enhances hematologic and immune recovery in mice and nonhuman primates.

Medications that can mitigate against radiation injury are limited. In this study, we investigated the ability of recombinant human growth hormone (rhGH) to mitigate against radiation injury in mice and nonhuman primates. BALB/c mice were irradiated with 7.5 Gy and treated post-irradiation with rhGH intravenously at a once daily dose of 20 microg/dose for 35 days. rhGH protected 17 out of 28 mice (60.7%) from lethal irradiation while only 3 out of 28 mice (10.7%) survived in the saline control group. A shorter course of 5 days of rhGH post-irradiation produced similar results. Compared with the saline control group, treatment with rhGH on irradiated BALB/c mice significantly accelerated overall hematopoietic recovery. Specifically, the recovery of total white cells, CD4 and CD8 T cell subsets, B cells, NK cells and especially platelets post radiation exposure were significantly accelerated in the rhGH-treated mice. Moreover, treatment with rhGH increased the frequency of hematopoietic stem/progenitor cells as measured by flow cytometry and colony forming unit assays in bone marrow harvested at day 14 after irradiation, suggesting the effects of rhGH are at the hematopoietic stem/progenitor level. rhGH mediated the hematopoietic effects primarily through their niches. Similar data with rhGH were also observed following 2 Gy sublethal irradiation of nonhuman primates. Our data demonstrate that rhGH promotes hematopoietic engraftment and immune recovery post the exposure of ionizing radiation and mitigates against the mortality from lethal irradiation even when administered after exposure.