Diet, psychosocial stress, and Alzheimer's disease-related neuroanatomy in female nonhuman primates.

INTRODUCTION: Associations between diet, psychosocial stress, and neurodegenerative disease, including Alzheimer's disease (AD), have been reported, but causal relationships are difficult to determine in human studies. METHODS: We used structural magnetic resonance imaging in a well-validated non-human primate model of AD-like neuropathology to examine the longitudinal effects of diet (Mediterranean vs Western) and social subordination stress on brain anatomy, including global volumes, cortical thicknesses and volumes, and 20 individual regions of interest (ROIs). RESULTS: Western diet resulted in greater cortical thicknesses, total brain volumes, and gray matter, and diminished cerebrospinal fluid and white matter volumes. Socially stressed subordinates had smaller whole brain volumes but larger ROIs relevant to AD than dominants. DISCUSSION: The observation of increased size of AD-related brain areas is consistent with similar reports of mid-life volume increases predicting increased AD risk later in life. While the biological mechanisms underlying the findings require future investigation, these observations suggest that Western diet and psychosocial stress instigate pathologic changes that increase risk of AD-associated neuropathology, whereas the Mediterranean diet may protect the brain.

Endometrium and endometriosis tissue mitochondrial energy metabolism in a nonhuman primate model.

BACKGROUND: Endometriosis is the growth of uterine lining (endometrium) outside of the uterus. In other chronic inflammatory diseases, mitochondrial dysfunction is suspected of playing a role in disease pathogenesis. However, little is known about endometriosis mitochondrial function or its effects on tissue metabolism. The objectives of this study were to analyze mitochondrial function in nonhuman primate (NHP) endometrium and endometriosis tissue and to identify the metabolic features of these tissues that may contribute to disease. METHODS: Mitochondrial function in endometriosis tissue and endometrium was measured using mitochondrial respirometry analysis to determine if changes in oxidative phosphorylation exist in endometrium and endometriosis tissue compared to control endometrium from clinically healthy NHPs. Targeted metabolomics and multidimensional statistical analysis were applied to quantify key metabolites in energy and amino acid biosynthesis pathways. RESULTS: Mitochondrial respirometry assays showed endometrium from NHPs with endometriosis had reduced complex II-mediated oxygen consumption rates (OCR) across all energy states (basal, p = 0.01; state 3, p = 0.02; state 3u, p = 0.04; state 4o, p = 0.008) and endometriosis tissue had reduced state 3, complex I-mediated OCR (p = 0.02) and respiratory control rates (p = 0.01) compared to normal endometrium. Targeted metabolomics performed on tissue revealed carnitine (p = 0.001), creatine phosphate (p = 0.01), NADH (p = 0.0001), FAD (p = 0.001), tryptophan (p = 0.0009), and malic acid (p = 0.005) were decreased in endometriosis tissue compared to normal endometrium samples. FAD (p = 0.004), tryptophan (p = 0.0004) and malic acid (p = 0.03) were significantly decreased in endometrium from NHPs with endometriosis compared to normal endometrium. Significant metabolites identified in endometriosis and endometrium samples from animals with endometriosis were part of amino acid biosynthesis or energy metabolism pathways. CONCLUSIONS: Here, endometrial mitochondrial energy production and metabolism were decreased in endometrium and endometriosis tissue. Decreased mitochondrial energy production may be due to oxidative stress-induced damage to mitochondrial DNA or membranes, a shift in cell metabolism, or decreased energy substrate; however, the exact cause remains unknown. Additional research is needed to determine the implications of reduced mitochondrial energy production and metabolism on endometriosis and endometrium.

Skeletal Muscle Mitochondrial Respiration Is Elevated in Female Cynomolgus Macaques Fed a Western Compared with a Mediterranean Diet.

BACKGROUND: Western diets are associated with increased incidences of obesity, hypertension, diabetes, and hypercholesterolemia, whereas Mediterranean diets, richer in polyphenols, monounsaturated fats, fruits, vegetables, poultry, and fish, appear to have cardiometabolic health benefits. Previous work has included population-based studies with limited evidence for causation or animal studies focused on single macro- or micronutrients; therefore, primate animal models provide an opportunity to determine potential mechanisms underlying the effects of dietary patterns on health and disease. OBJECTIVE: The aim of this study was to determine the effects of whole dietary patterns, either a Western or Mediterranean diet, on skeletal muscle mitochondrial bioenergetics in cynomolgus macaques. METHODS: In this study, 22 adult female cynomolgus macaques (∼11-14 y by dentition) were fed either a Western or Mediterranean diet for 30 mo. The Western diet was designed to mimic the diet of a middle-aged American woman and the Mediterranean diet included key aspects of Mediterranean diets studied in humans, such as plant-based proteins and fat, complex carbohydrates, and fiber. Diets were matched on macronutrient composition (16% protein, 54% carbohydrate, and 31% fat) and cholesterol content. Skeletal muscle was collected for high-resolution respirometry, citrate synthase activity, and western blot measurements. Pearson correlation analysis between respirometry measures and measures of carbohydrate metabolism was also performed. RESULTS: We found that consumption of a Western diet resulted in significantly higher mitochondrial respiration with fatty acid oxidation (FAO) (53%), FAO + complex I (52%), complex I + II (31%), max electron transport system (ETS) (31%), and ETS rotenone sensitive (31%) than did consumption of a Mediterranean diet. In addition, measures of respiration in response to fatty acids were significantly and positively correlated with both insulin resistance and plasma insulin concentrations. CONCLUSIONS: This study highlights the importance of dietary composition in mitochondrial bioenergetics and that diet can influence skeletal muscle mitochondrial respiration independently of other factors such as macronutrient composition.

Novel fatty acyl apoE mimetic peptides have increased potency to reduce plasma cholesterol in mice and macaques.

Ac-hE18A-NH2 is a dual-domain apoE mimetic peptide that possesses the putative receptor binding domain from apoE (LRKLRKRLLR, denoted hE; residues 141-150) covalently attached to lipid-associating peptide 18A. Like apoE, Ac-hE18A-NH2 reduces plasma cholesterol in animal models and exhibits anti-inflammatory properties independent of its cholesterol-reducing effect. Ac-hE18A-NH2 has already undergone phase I clinical trials as a lipid-lowering agent. To explore the therapeutic potential more, we designed and synthesized new analogues by linking ɑ-aminohexanoic acid, octanoic acid, or myristic acid to LRRLRRRLLR-18A-NH2 ([R]hE18A-NH2) and examined the cholesterol-lowering potency in animals. The modified peptides effectively reduced plasma cholesterol in apoE-null mice fed standard chow or a Western diet; the myristyl analogue was the most effective. A single administration of the myristyl analogue reduced plasma total and LDL cholesterol in a dose-dependent manner in hypercholesterolemic cynomolgus macaques for up to 1 week despite the continuation of a cholesterol-supplemented diet. The myristyl peptide (7.4 mg/kg) reduced total and LDL cholesterol at 24 h by 64% and 74%, respectively; plasma HDL levels were modestly reduced and returned to baseline by day 7. These new analogues should exhibit enhanced potency at lower doses than Ac-hE18A-NH2, which may make them attractive therapeutic candidates for clinical trials.

Effects of long-term sertraline treatment and depression on coronary artery atherosclerosis in premenopausal female primates.

OBJECTIVES: Major depressive disorder and coronary heart disease often co-occur in the same individuals. Selective serotonin reuptake inhibitors (SSRIs) are widely prescribed for depression and other disorders, but their effects on coronary heart disease risk remain unclear. We determined the effects of an SSRI on coronary artery atherosclerosis (CAA) in an established nonhuman primate model used to clarify the association between depression and CAA. METHODS: Forty-two adult female cynomolgus macaques consuming a Western diet were characterized during an 18-month pretreatment phase and assigned to SSRI (sertraline hydrochloride 20 mg/kg, per os, once a day) or placebo balanced on pretreatment depression, body weight (BW), and iliac artery atherosclerosis extent measured via biopsy. After 18 months, CAA extent was measured using histomorphometry. RESULTS: Before and during treatment, depressed monkeys had lower BW, body mass index, and plasma high-density lipoprotein cholesterol, and higher heart rates during the pretreatment (p < .01) but not the treatment phase (p = .17). There were no pretreatment differences between the sertraline and placebo groups. Sertraline reduced anxious behavior but had no effect on BW, body mass index, heart rate, plasma lipids, or depression. CAA, analyzed by a 2 (depressed, nondepressed) × 2 (placebo, sertraline) × 3 (coronary arteries) analysis of covariance adjusted for pretreatment iliac atherosclerosis, was greater in depressed than in nondepressed monkeys (p < .036), and in sertraline than in placebo-treated monkeys (p = .040). The observed CAA extent in depressed monkeys treated with sertraline was 4.9 times higher than that in untreated depressed monkeys, and 6.5 times higher than that in nondepressed monkeys, on average. CONCLUSIONS: Depressed animals developed more CAA, and long-term treatment with sertraline resulted in more extensive CAA.

Differential effects of western versus mediterranean diets and psychosocial stress on ovarian function in female monkeys (Macaca fascicularis).

Ovarian dysfunction increases risk for chronic diseases of aging including cardiovascular disease, depression, cognitive impairment, as well as bone and muscle loss which promote frailty. Psychosocial stress can disrupt ovarian function, and recent observations suggest that consumption of a Western Diet may also. Determination of causal relationships among diet, psychosocial stress, and ovarian physiology is difficult in humans. Long-tailed (a.k.a. cynomolgus) macaques (Macaca fascicularis) are an excellent translational model for the study of diet and psychosocial effects on ovarian physiology and aging-related processes. They have 28-day menstrual cycles with hormonal fluctuations like those of women, and similar physiologic responses to alterations and/or cessation of cyclicity. We examined ovarian function in 38 middle-aged socially housed females fed either a Western or Mediterranean diet for 31 months (≈ a 9-year period for humans). During the last year, we examined cycle length and peak progesterone per cycle using blood sampling (3/week) and vaginal swabbing for menses (6/week). Repeated measures analysis revealed a circannual pattern consistent with increased menstrual cycle disturbance during the late Summer and early Fall (F(11,348)= 4.05 p < 0.001). In addition, both Western diet (F(1,34)= 3.99; p = 0.05) and the stress of low social status (F(1,34)= 3.99; p = 0.04) reduced mean progesterone levels. Thus, on average, subordinates in the Western group had the lowest average progesterone levels (10.02 ng/pl). Compared to Western diets, Mediterranean diets exhibited protective effects via menstrual cycle regularity. For dominant monkeys, consuming Mediterranean diets resulted in significantly greater likelihood of having regular menstrual cycles. Mediterranean diets also protected individuals from shorter than normal menstrual cycles. The relationships between diet and menstrual regularity were partially mediated by both adrenal reactivity and social isolation. This study demonstrates the additive negative effects of poor diet and psychosocial stress on ovarian physiology in mid-life and lays the groundwork for future investigations to uncover their impact on metabolic signatures of accelerated aging. The results also suggest that - compared to Western-style diets - a Mediterranean diet may exert a protective influence against ovarian dysfunction and its pathologic sequelae.

Contrast enhanced computed tomography of small ruminants: Caprine and ovine.

The use of small ruminants, mainly sheep and goats, is increasing in biomedical research. Small ruminants are a desirable animal model due to their human-like anatomy and physiology. However, the large variability between studies and lack of baseline data on these animals creates a barrier to further research. This knowledge gap includes a lack of computed tomography (CT) scans for healthy subjects. Full body, contrast enhanced CT scans of caprine and ovine subjects were acquired for subsequent modeling studies. Scans were acquired from an ovine specimen (male, Khatadin, 30-35 kg) and caprine specimen (female, Nubian 30-35 kg). Scans were acquired with and without contrast. Contrast enhanced scans utilized 1.7 mL/kg of contrast administered at 2 mL/s and scans were acquired 20 seconds, 80 seconds, and 5 minutes post-contrast. Scans were taken at 100 kV and 400 mA. Each scan was reconstructed using a bone window and a soft tissue window. Sixteen full body image data sets are presented (2 specimens by 4 contrast levels by 2 reconstruction windows) and are available for download through the form located at: https://redcap.link/COScanData. Scans showed that the post-contrast timing and scan reconstruction method affected structural visualization. The data are intended for further biomedical research on ruminants related to computational model development, device prototyping, comparative diagnostics, intervention planning, and other forms of translational research.

Psychosocial stress increases risk for type 2 diabetes in female cynomolgus macaques consuming a western diet.

Chronic psychosocial stress is associated with increased risk of many chronic diseases including type 2 diabetes mellitus. However, it is difficult to establish a causal relationship between stress and diabetes in human studies because stressors often are self-reported and may be distant in time from metabolic consequences. Macaques are useful models of the effects of chronic psychosocial stress on health and may develop obesity and diabetes similar to human beings. Thus, we studied the relationships between social subordination stress - a well-validated psychological stressor in macaques - and body composition and carbohydrate metabolism in socially housed, middle-aged female cynomolgus monkeys (Macaca fascicularis; n = 42). Following an 8-week baseline phase, the monkeys were fed a Western diet for 36 months (about equivalent to 10 human years). Social status was determined based on the outcomes of agonistic interactions (X¯= 33.3 observation hours/monkey). Phenotypes collected included plasma cortisol, body composition, circulating markers of glucose metabolism, activity levels, and heart rate variability measured as RMSSD (root of mean square of successive differences) and SDDN (standard deviation of beat to beat interval) after 1.5- and 3-years on diet. Mixed model analyses of variance revealed that aggression received, submissions sent, and cortisol were higher, and RMSSD and SDNN were lower in subordinates than dominants (social status: p < 0.05). After 3 years of Western diet consumption, fasting triglyceride, glucose and insulin concentrations, calculated insulin resistance (HOMA-IR), body weight and body fat mass increased in all animals (time: all p's < 0.05); however, the increase in fasting glucose and HOMA-IR was significantly greater in subordinates than dominants (time x social status: p's < 0.05). Impaired glucose metabolism, (glucose > 100 mg/dl) incidence was significantly higher in subordinates (23%) than dominants (0%) (Fisher's exact test, p < 0.05). These findings suggest that chronic psychosocial stress, on a Western diet background, significantly increases type 2 diabetes risk in middle-aged female primates.

Pharmacological Profile and Ocular Hypotensive Effects of Cromakalim Prodrug 1, a Novel ATP-Sensitive Potassium Channel Opener, in Normotensive Dogs and Nonhuman Primates.

Purpose: To evaluate pharmacokinetic parameters and ocular hypotensive effects of cromakalim prodrug 1 (CKLP1) in normotensive large animal models. Methods: Optimal CKLP1 concentration was determined by dose response and utilized in short- (5-8 days) and long-term (60 days) evaluation in hound dogs (n = 5) and African Green Monkeys (n = 5). Blood pressure was recorded 3-5 times per week with a tail cuff. Concentrations of CKLP1 and the parent compound levcromakalim were assessed in hound dog plasma and select tissues by LC-MS/MS after bilateral ocular treatment with CKLP1 for 8 days. Pharmacokinetic parameters were calculated from days 1, 4, and 8 data. After necropsy, histology was assessed in 43 tissue samples from each animal. Results: In hound dogs and African Green monkeys, 10 mM CKLP1 (optimal concentration) significantly lowered intraocular pressure (IOP) by 18.9% ± 1.1% and 16.7% ± 6.7%, respectively, compared with control eyes (P < 0.05). During treatment, no significant change in systolic or diastolic blood pressure was observed in either species (P > 0.1). Average values for half-life of CKLP1 was 295.3 ± 140.4 min, Cmax, 10.5 ± 1.6 ng/mL, and area under the concentration vs. time curve (AUClast) 5261.4 ± 918.9 ng·min/mL. For levcromakalim, average values of half-life were 96.2 ± 27 min, Cmax 1.2 ± 0.2 ng/mL, and AUClast 281.2 ± 110.8 ng·min/mL. No significant pathology was identified. Conclusions: CKLP1 lowered IOP in hound dogs and African green monkeys with no effect on systemic blood pressure. Ocular topical treatment of CKLP1 showed excellent tolerability even after extended treatment periods.

Contrasting effects of Western vs Mediterranean diets on monocyte inflammatory gene expression and social behavior in a primate model.

Dietary changes associated with industrialization increase the prevalence of chronic diseases, such as obesity, type II diabetes, and cardiovascular disease. This relationship is often attributed to an 'evolutionary mismatch' between human physiology and modern nutritional environments. Western diets enriched with foods that were scarce throughout human evolutionary history (e.g. simple sugars and saturated fats) promote inflammation and disease relative to diets more akin to ancestral human hunter-gatherer diets, such as a Mediterranean diet. Peripheral blood monocytes, precursors to macrophages and important mediators of innate immunity and inflammation, are sensitive to the environment and may represent a critical intermediate in the pathway linking diet to disease. We evaluated the effects of 15 months of whole diet manipulations mimicking Western or Mediterranean diet patterns on monocyte polarization in a well-established model of human health, the cynomolgus macaque (Macaca fascicularis). Monocyte transcriptional profiles differed markedly between diets, with 40% of transcripts showing differential expression (FDR < 0.05). Monocytes from Western diet consumers were polarized toward a more proinflammatory phenotype. The Western diet shifted the co-expression of 445 gene pairs, including small RNAs and transcription factors associated with metabolism and adiposity in humans, and dramatically altered behavior. For example, Western-fed individuals were more anxious and less socially integrated. These behavioral changes were also associated with some of the effects of diet on gene expression, suggesting an interaction between diet, central nervous system activity, and monocyte gene expression. This study provides new molecular insights into an evolutionary mismatch and uncovers new pathways through which Western diets alter monocyte polarization toward a proinflammatory phenotype.

Diet, obesity, and the gut microbiome as determinants modulating metabolic outcomes in a non-human primate model.

BACKGROUND: The objective of this study was to increase understanding of the complex interactions between diet, obesity, and the gut microbiome of adult female non-human primates (NHPs). Subjects consumed either a Western (n=15) or Mediterranean (n=14) diet designed to represent human dietary patterns for 31 months. Body composition was determined using CT, fecal samples were collected, and shotgun metagenomic sequencing was performed. Gut microbiome results were grouped by diet and adiposity. RESULTS: Diet was the main contributor to gut microbiome bacterial diversity. Adiposity within each diet was associated with subtle shifts in the proportional abundance of several taxa. Mediterranean diet-fed NHPs with lower body fat had a greater proportion of Lactobacillus animalis than their higher body fat counterparts. Higher body fat Western diet-fed NHPs had more Ruminococcus champaneliensis and less Bacteroides uniformis than their low body fat counterparts. Western diet-fed NHPs had significantly higher levels of Prevotella copri than Mediterranean diet NHPs. Western diet-fed subjects were stratified by P. copri abundance (P. copriHIGH versus P. copriLOW), which was not associated with adiposity. Overall, Western diet-fed animals in the P. copriHIGH group showed greater proportional abundance of B. ovatus, B. faecis, P. stercorea, P. brevis, and Faecalibacterium prausnitzii than those in the Western P. copriLOW group. Western diet P. copriLOW subjects had a greater proportion of Eubacterium siraeum. E. siraeum negatively correlated with P. copri proportional abundance regardless of dietary consumption. In the Western diet group, Shannon diversity was significantly higher in P. copriLOW when compared to P. copriHIGH subjects. Furthermore, gut E. siraeum abundance positively correlated with HDL plasma cholesterol indicating that those in the P. copriLOW population may represent a more metabolically healthy population. Untargeted metabolomics on urine and plasma from Western diet-fed P. copriHIGH and P. copriLOW subjects suggest early kidney dysfunction in Western diet-fed P. copriHIGH subjects. CONCLUSIONS: In summary, the data indicate diet to be the major influencer of gut bacterial diversity. However, diet and adiposity must be considered together when analyzing changes in abundance of specific bacterial taxa. Interestingly, P. copri appears to mediate metabolic dysfunction in Western diet-fed NHPs. Video abstract.

Validation of multi-detector computed tomography as a non-invasive method for measuring ovarian volume in macaques (Macaca fascicularis).

The purpose of this study was to validate low radiation dose, contrast-enhanced, multi-detector computed tomography (MDCT) as a non-invasive method for measuring ovarian volume in macaques. Computed tomography scans of four known-volume phantoms and nine mature female cynomolgus macaques were acquired using a previously described, low radiation dose scanning protocol, intravenous contrast enhancement, and a 32-slice MDCT scanner. Immediately following MDCT, ovaries were surgically removed and the ovarian weights were measured. The ovarian volumes were determined using water displacement. A veterinary radiologist who was unaware of actual volumes measured ovarian CT volumes three times, using a laptop computer, pen display tablet, hand-traced regions of interest, and free image analysis software. A statistician selected and performed all tests comparing the actual and CT data. Ovaries were successfully located in all MDCT scans. The iliac arteries and veins, uterus, fallopian tubes, cervix, ureters, urinary bladder, rectum, and colon were also consistently visualized. Large antral follicles were detected in six ovaries. Phantom mean CT volume was 0.702+/-SD 0.504 cc and the mean actual volume was 0.743+/-SD 0.526 cc. Ovary mean CT volume was 0.258+/-SD 0.159 cc and mean water displacement volume was 0.257+/-SD 0.145 cc. For phantoms, the mean coefficient of variation for CT volumes was 2.5%. For ovaries, the least squares mean coefficient of variation for CT volumes was 5.4%. The ovarian CT volume was significantly associated with actual ovarian volume (ICC coefficient 0.79, regression coefficient 0.5, P=0.0006) and the actual ovarian weight (ICC coefficient 0.62, regression coefficient 0.6, P=0.015). There was no association between the CT volume accuracy and mean ovarian CT density (degree of intravenous contrast enhancement), and there was no proportional or fixed bias in the CT volume measurements. Findings from this study indicate that MDCT is a valid non-invasive technique for measuring the ovarian volume in macaques.

Mediterranean diet, stress resilience, and aging in nonhuman primates.

Persistent psychological stress increases the risk of many chronic diseases of aging. Little progress has been made to effectively reduce stress responses or mitigate stress effects suggesting a need for better understanding of factors that influence stress responses. Limited evidence suggests that diet may be a factor in modifying the effects of stress. However, long-term studies of diet effects on stress reactive systems are not available, and controlled randomized clinical trials are difficult and costly. Here we report the outcomes of a controlled, randomized preclinical trial of the effects of long-term consumption (31 months, ~ equivalent to 9 human years) of Western versus Mediterranean - like diets on behavioral and physiological responses to acute (brief social separation) and chronic (social subordination) psychosocial stress in 38 adult, socially-housed, female cynomolgus macaques. Compared to animals fed a Western diet, those fed the Mediterranean diet exhibited enhanced stress resilience as indicated by lower sympathetic activity, brisker and more overt heart rate responses to acute stress, more rapid recovery, and lower cortisol responses to acute psychological stress and adrenocorticotropin (ACTH) challenge. Furthermore, age-related increases in sympathetic activity and cortisol responses to stress were delayed by the Mediterranean diet. Population level diet modification in humans has been shown to be feasible. Our findings suggest that population-wide adoption of a Mediterranean-like diet pattern may provide a cost-effective intervention on psychological stress and promote healthy aging with the potential for widespread efficacy.

Effects of 4-vinylcyclohexene diepoxide on peripubertal and adult Sprague-Dawley rats: ovarian, clinical, and pathologic outcomes.

Young rats treated daily with intraperitoneal 4-vinylcyclohexene diepoxide (VCD) undergo selective destruction of primordial follicles, resulting in gradual ovarian failure resembling the menopausal transition in women. To determine whether VCD has similar effects on ovaries of older rats, adult and peripubertal Sprague-Dawley rats were injected intraperitoneally daily for 30 d with vehicle or VCD at 40 or 80 mg/kg. Body weight, food intake, complete blood counts, and markers of liver injury and renal function were measured during VCD treatment. Complete gross necropsy and microscopic observations were performed on day 31, and ovarian follicles were counted. At 80 mg/kg, VCD destroyed primordial and primary follicles to a similar extent in both adult and peripubertal animals, although adult rats likely started with fewer follicles and therefore approached follicle depletion. Treatment with VCD did not affect body weight, but food intake was reduced in both adult and peripubertal rats treated with 80 mg/kg VCD. Adult rats treated with 80 mg/kg VCD had neutrophilia and increased BUN and creatinine; in addition, 4 of these rats were euthanized on days 25 or 26 due to peritonitis. VCD treatment did not increase alanine aminotransferase levels, a marker of liver injury, although the 80-mg/kg dose increased liver weights. In conclusion, VCD effectively destroys small preantral follicles in adult Sprague-Dawley rats, making them a suitable model of the menopausal transition of women. However, because adult rats were more sensitive to the irritant properties of VCD, the use of a lower dose should be considered.

Mediterranean versus Western Diet Effects on Caloric Intake, Obesity, Metabolism, and Hepatosteatosis in Nonhuman Primates.

OBJECTIVE: This study aimed to determine the effects of humanlike Western and Mediterranean diets on caloric intake, obesity, metabolism, and hepatosteatosis in an established nonhuman primate model of obesity, cardiometabolic syndrome, type 2 diabetes, and atherosclerosis. METHODS: A 38-month, randomized, preclinical, nonhuman primate primary prevention trial of 38 socially housed, middle-aged adult females was conducted. The monkeys were characterized during a 7-month baseline phase while consuming chow and then randomized to either Western or Mediterranean diets; the groups were balanced on baseline characteristics. Western and Mediterranean diets were formulated to closely reflect human diets, matched on macronutrient content, with protein and fat derived largely from animal sources in the Western diet and plant sources in the Mediterranean diet. Food consumption, activity levels, energy expenditure, body composition, carbohydrate metabolism, and hepatosteatosis were measured during baseline and treatment phases. RESULTS: The Western diet increased caloric intake for the first 6 months and body fat, activity, energy expenditure, insulin resistance, and hepatosteatosis after 2.5 years, whereas the Mediterranean diet reduced triglyceride levels. CONCLUSIONS: This is the first report of differential caloric intake and obesity with long-term consumption of a Western versus Mediterranean diet under controlled experimental conditions and the first experimental evidence that a Mediterranean diet protects against hepatosteatosis compared with a Western diet.

Both diet and Helicobacter pylori infection contribute to atherosclerosis in pre- and postmenopausal cynomolgus monkeys.

A number of viruses and bacterial species have been implicated as contributors to atherosclerosis, potentially providing novel pathways for prevention. Epidemiological studies examining the association between Helicobacter pylori and cardiovascular disease have yielded variable results and no studies have been conducted in nonhuman primates. In this investigation, we examined the relationship between H. pylori infection and atherosclerosis development in socially housed, pre- and postmenopausal cynomolgus macaques consuming human-like diets. Ninety-four premenopausal cynomolgus monkeys (Macaca fascicularis) were fed for 36 months an atherogenic diet deriving its protein from either casein lactalbumin(CL) or high isoflavone soy (SOY). Animals were then ovariectomized and fed either the same or the alternate diet for an additional 36 months. Iliac artery biopsies were obtained at the time of ovariectomy and iliac and coronary artery sections were examined at the end of the study. Evidence of H. pylori infection was found in 64% of the monkeys and 46% of animals had live H. pylori within coronary atheromas as determined by mRNA-specific in situ hybridization. There was a significant linear relationship between the densities of gastric and atheroma organisms. Helicobactor pylori infection correlated with increased intimal plaque area and thickness at both the premenopausal and postmenopausal time points and regardless of diet (p< 0.01), although animals consuming the SOY diet throughout had the least amount of atherosclerosis. Additionally, plasma lipid profiles, intimal collagen accumulation, ICAM-1, and plaque macrophage densities were adversely affected by H. pylori infection among animals consuming the CL diet, while the SOY diet had the opposite effect. Plaque measurements were more highly associated with the densities of cagA-positive H. pylori within coronary atheromas than with the densities of gastric organisms, whereas plasma lipid changes were associated with H. pylori infection, but not cagA status. This study provides strong evidence that live H. pylori infects atheromas, exacerbates atherosclerotic plaque development, and alters plasma lipid profiles independently of diet or hormonal status. Finally, socially subordinate animals relative to their dominant counterparts had a greater prevalence of H. pylori, suggesting a stress effect. The results indicate that early H. pylori eradication could prevent or delay development of cardiovascular disease.

Estrogen effects on epithelial proliferation and benign proliferative lesions in the postmenopausal primate mammary gland.

Proliferative lesions of the mammary gland are risk markers and potential precursors for the development of breast cancer in postmenopausal women. In this study we evaluated mammary epithelial proliferation and proliferative lesions in a group of 63 aged postmenopausal macaques randomized by social group to receive one of three experimental diets for 8 months: (1) control; (2) control with 17beta-estradiol (E2) at the human equivalent dose of 1.0 mg per day; and (3) control with the soy phytoestrogen equol (EQ) at the human equivalent dose of 105 mg per day. In normal mammary epithelium, treatment with E2 but not EQ resulted in greater proliferation, epithelial area, and progesterone receptor expression (P<0.05 for all). Mammary lesions included columnar cell change (26/63), columnar cell hyperplasia with and without atypia (13/63), atypical ductal hyperplasia (6/63), and atypical lobular hyperplasia (3/63). Lesions were most common within terminal ductal lobular units. The prevalence of columnar cell hyperplasia (total and atypical cases) was higher in animals treated with E2 compared to control (P<0.05 for both). Compared to normal mammary epithelium, columnar cell lesions (CCLs) showed greater constitutive expression of estrogen receptor-alpha across all groups (P<0.001) and greater expression of progesterone receptor in response to E2 (P<0.01). Independent of treatment, animals with CCLs on histology had greater gene expression of estrogen receptor-alpha and markers of estrogen receptor activity (trefoil factor 1) and proliferation (gene for Ki67 antigen) at a site contralateral to the CCL (P<0.05 for all). These findings demonstrate that the terminal ductal lobular units of the postmenopausal mammary gland contain morphologically distinct cell populations that may hyperrespond to E2 exposure, resulting in specific types of hyperplastic lesions.

Soy-tibolone combination - effect on lipids in postmenopausal monkeys and women.

OBJECTIVES: To determine whether co-administration of soy during tibolone treatment would prevent tibolone-induced dyslipoproteinemia in postmenopausal monkeys and women. METHODS: Surgically postmenopausal cynomolgus monkeys (n = 18) were assigned randomly to one of four dietary regimens in a Latin Square crossover design, such that all animals received all diets for 14 weeks with a 4-week washout period: (1) casein/lactalbumin (CL); (2) tibolone (Tib, 1.25 mg/day women's equivalent); (3) soy (138 mg isoflavones/day women's equivalent); (4) Soy + Tib. Postmenopausal women on tibolone treatment were randomized to receive soy powder (52 g of soy protein containing 112 mg isoflavones) or placebo (containing 52 g of milk protein) daily in a crossover trial for 8 weeks with a 4-week washout period. RESULTS: Monkeys given Tib alone had approximately 14% increase in plasma LDL + VLDL-C; whereas those given soy combined with tibolone had significant ( approximately 22%) reductions. Tib treated monkeys had reductions in plasma HDL-C of about 48% vs. no reductions in Soy + Tib. In postmenopausal women using tibolone, soy reduced plasma LDL-C concentrations by approximately 10% from baseline without a change in HDL-C. CONCLUSIONS: Co-administration of soy during tibolone treatment improved the lipoprotein profile in both monkeys and women; however, the effects were more robust in monkeys.

Consumption of Mediterranean versus Western Diet Leads to Distinct Mammary Gland Microbiome Populations.

Recent identification of a mammary gland-specific microbiome led to studies investigating bacteria populations in breast cancer. Malignant breast tumors have lower Lactobacillus abundance compared with benign lesions, implicating Lactobacillus as a negative regulator of breast cancer. Diet is a main determinant of gut microbial diversity. Whether diet affects breast microbiome populations is unknown. In a non-human primate model, we found that consumption of a Western or Mediterranean diet modulated mammary gland microbiota and metabolite profiles. Mediterranean diet consumption led to increased mammary gland Lactobacillus abundance compared with Western diet-fed monkeys. Moreover, mammary glands from Mediterranean diet-fed monkeys had higher levels of bile acid metabolites and increased bacterial-processed bioactive compounds. These data suggest that diet directly influences microbiome populations outside the intestinal tract in distal sites such as the mammary gland. Our study demonstrates that diet affects the mammary gland microbiome, establishing an alternative mechanistic pathway for breast cancer prevention.

Systemic Iron Deficiency in a Nonhuman Primate Model of Endometriosis.

Endometriosis is characterized by endometrial tissue development outside the uterus. Anemia and iron depletion do not commonly accompany endometriosis in women, despite chronic abdominal inflammation and heavy menstrual bleeding. The objective of this study was to examine iron kinetics associated with endometriosis by using a NHP model, to better understand the underlying mechanism of abnormal hematogram values in women with endometriosis. Hematologic data from 46 macaques with endometriosis were examined for signs of iron depletion. Bone marrow, liver, and serum were used to elucidate whether iron loss or inflammation best explained the hematologic findings. Additional serum markers and intestinal biopsies from NHP with and without endometriosis were evaluated for patterns in iron kinetics across the menstrual cycle and for relative dietary iron-absorbing capacity. Almost half of the NHP with endometriosis were anemic. Overall, NHP had decreased RBC counts, increased MCV, increased percentage of reticulocytes, decreased serum hepcidin, and decreased hepatic and bone marrow iron. Intestinal expression of ferroportin 1, a mediator of iron absorption, was increased, indicating that despite high dietary iron, intestinal iron absorption did not compensate for iron losses. We concluded that use of oral iron supplementation alone does not replenish iron stores in endometriosis. Consequently, iron stores should be evaluated in women with endometriosis, even without overt clinical signs of anemia.