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.

Macrophage Phenotypes and Gene Expression Patterns Are Unique in Naturally Occurring Metabolically Healthy Obesity.

Obesity impacts 650 million individuals globally, often co-occurring with metabolic syndrome. Though many obese individuals experience metabolic abnormalities (metabolically unhealthy obese [MUO]), ~30% do not (metabolically healthy obese [MHO]). Conversely, >10% of lean individuals are metabolically unhealthy (MUL). To evaluate the physiologic drivers of these phenotypes, a 44-animal African green monkey cohort was selected using metabolic syndrome risk criteria to represent these four clinically defined health groups. Body composition imaging and subcutaneous adipose tissue (SQ AT) biopsies were collected. Differences in adipocyte size, macrophage subtype distribution, gene expression, vascularity and fibrosis were analyzed using digital immunohistopathology, unbiased RNA-seq, endothelial CD31, and Masson's trichrome staining, respectively. MHO AT demonstrated significant increases in M2 macrophages (p = 0.02) and upregulation of fatty acid oxidation-related terms and transcripts, including FABP7 (p = 0.01). MUO AT demonstrated downregulation of these factors and co-occurring upregulation of immune responses. These changes occurred without differences in AT distributions, adipocyte size, AT endothelial cells, collagen I deposition, or circulating cytokine levels. Without unhealthy diet consumption, healthy obesity is defined by an increased SQ AT M2/M1 macrophage ratio and lipid handling gene expression. We highlight M2 macrophages and fatty acid oxidation as targets for improving metabolic health with obesity.

Visceral adipose microbial and inflammatory signatures in metabolically healthy and unhealthy nonhuman primates.

OBJECTIVE: Obesity is a key risk factor for metabolic syndrome (MetS); however, >10% of lean individuals meet MetS criteria. Visceral adipose tissue (VAT) disproportionately contributes to inflammation and insulin resistance compared with subcutaneous fat depots. The primary aim of this study was to profile tissue microbiome components in VAT over a wide range of metabolic statuses in a highly clinically relevant model. METHODS: VAT was profiled from nonhuman primates that naturally demonstrate four distinct health phenotypes despite consuming a healthy diet, namely metabolically healthy lean and obese and metabolically unhealthy lean and obese. RESULTS: VAT biopsied from unhealthy lean and obese nonhuman primates demonstrated upregulation of immune signaling pathways, a tissue microbiome enriched in gram-negative bacteria including Pseudomonas, and deficiencies in anti-inflammatory adipose tissue M2 macrophages. VAT microbiomes were distinct from fecal microbiomes, and fecal microbiomes did not differ by metabolic health group, which was in contrast to the VAT bacterial communities. CONCLUSIONS: Immune activation with gram-negative VAT microbial communities is a consistent feature in elevated MetS risk in both lean and obesity states.

Replacement substance P reduces cardiac fibrosis in monkeys with type 2 diabetes.

BACKGROUND: Type 2 diabetes mellitus (T2DM)-associated cardiac fibrosis contributes to heart failure. We previously showed that diabetic mice with cardiomyopathy, including cardiac fibrosis, exhibit low levels of the neuropeptide substance P; exogenous replacement of substance P reversed cardiac fibrosis, independent of body weight, blood glucose and blood pressure. We sought to elucidate the effectiveness and safety of replacement substance P to ameliorate or reverse cardiac fibrosis in type 2 diabetic monkeys. METHODS: Four female T2DM African Green monkeys receive substance P (0.5 mg/Kg/day S.Q. injection) for 8 weeks. We obtained cardiac magnetic resonance imaging and blood samples to assess left ventricular function and fibrosis by T1 map-derived extracellular volume as well as circulating procollagen type I C-terminal propeptide. Hematological parameters for toxicities were also assessed in these monkeys and compared with three female T2DM monkeys receiving saline S.Q. as a safety comparison group. RESULTS: Diabetic monkeys receiving replacement substance P exhibited a ∼20% decrease in extracellular volume (p = 0.01), concomitant with ∼25% decrease procollagen type I C-terminal propeptide levels (p = 0.008). Left ventricular ejection fraction was unchanged with substance P (p = 0.42); however, circumferential strain was improved (p < 0.01). Complete blood counts, glycosylated hemoglobin A1c, lipids, liver and pancreatic enzymes, and inflammation markers were unchanged (p > 0.05). CONCLUSIONS: Replacement substance P reversed cardiac fibrosis in a large preclinical model of type 2 diabetes, independent of glycemic control. No hematological or organ-related toxicity was associated with replacement substance P. These results strongly support a potential application for replacement substance P as safe therapy for diabetic cardiac fibrosis.

Xenon-enhanced computed tomography assessment of brown adipose tissue distribution and perfusion in lean, obese, and diabetic primates.

OBJECTIVE: This study aimed to validate xenon-enhanced computed tomography (XECT) for the detection of brown adipose tissue (BAT) and to use XECT to assess differences in BAT distribution and perfusion between lean, obese, and diabetic nonhuman primates (NHPs). METHODS: Whole-body XECT imaging was performed in anesthetized rhesus and vervet monkeys during adrenergic stimulation of BAT thermogenesis. In XECT images, BAT was identified as fat tissue that, during xenon inhalation, underwent significant radiodensity enhancement compared with subcutaneous fat. To measure BAT blood flow, BAT radiodensity enhancement was measured over time on the six computed tomography scans acquired during xenon inhalation. Postmortem immunohistochemical staining was used to confirm imaging findings. RESULTS: XECT was able to correctly identify all BAT depots that were confirmed at necropsy, enabling construction of the first comprehensive anatomical map of BAT in NHPs. A significant decrease in BAT perfusion was found in diabetic animals compared with obese animals and healthy animals, as well as absence of axillary BAT and significant reduction of supraclavicular BAT in diabetic animals compared with obese and lean animals. CONCLUSIONS: The use of XECT in NHP models of obesity and diabetes allows the analysis of the impact of metabolic status on BAT mass and perfusion.

Biomarkers of senescence in non-human primate adipose depots relate to aging.

Forty-three female African green monkeys (Chlorocebus aethiops sabaeus) were selected to represent young adult to advanced geriatric ages (7-24 years) to exhibit a wide range of obesity status (8-53% body fat) and diverse metabolic syndrome criteria such as diabetes, dyslipidemia, and hypertension. Subcutaneous and visceral adipose tissues were collected and evaluated for the presence of senescence cells in both whole tissue and single-cell isolates from subcutaneous sources, utilizing senescence-associated ß-galactosidase (SAß-gal) staining. Plasma samples were analyzed for selected metabolic and inflammatory biomarkers related to the senescence-associated secretory profile. Our results indicated that tissue staining scores did not differ between subcutaneous and intra-abdominal visceral depots and were highly related within individuals. Tissue staining was significantly associated with chronological age; however, no associations with fatness or metabolic syndrome criteria were observed. Associations with age were unchanged when obesity status was included in regression models. Isolated cell staining did positively relate to age but not tissue staining, suggesting some of the SAß-gal-positive cells were stromal vascular cells or small adipocytes, but that mature large adipocytes, filtered out in the cell isolation process, are also likely to exhibit positive SAß-gal staining. Plasminogen activator inhibitor-1 (PAI-1) concentration in circulation was the sole inflammation-related biomarker that positively associated with age and is considered to be a marker of senescent cell burden. Our study is the largest, most comprehensive assessment of adipose SAß-gal staining in a relevant animal model of human aging, and confirms that this senescence-associated biomarker specifically indicates an age-related process.