Following collection of baseline data, 16 monkeys had been assigned to an atherogenic diet and 7 monkeys to a control diet. We had quarterly measured over a two-year period: blood pressure, body weight, and morphometric measurements, blood pressure, and lipid profile. Preliminary statistical analyses showed that at baseline, cholesterol (p=0.39), triglycerides (p=0.64), and glucose (p=0.18) were not different between the control and treated groups. At the second follow-up, cholesterol levels in the treated group were different from controls (p<0.0001). The correlation of these parameters with other covariates found that at baseline, cholesterol was negatively associated with body weight in the controls but not in treated. There was a borderline significance with SBP and DBP, again only in the controls. At the second follow-up, glucose was related with body weight in the controls but not in the treated group. For cholesterol it was noted that the treatment group with time interaction was significant, indicating that treatment on these monkeys over a period of time had an effect on cholesterol levels. In the glucose model;first, age was a significant predictor, while in the triglyceride model neither first age, nor time nor the treatment group and none of the interactions were significant. In addition to the blood chemistry, four imaging parameters, namely contrast enhancement, wall thickness, lumen diameter and T2thickest triad, will be analyzed at the same time points as the other measures were done and correlate these parameters with the blood parameters to find associations at all time points. After sacrifice, we harvested arterial tissue. Histochemical observation and morphological analysis indicated that age increases intimal thickness and medial thickness along with atherosclerotic lesions in these domestic monkeys fed normal diets. Interestingly, age affects fat deposition within arterial walls in these domestic monkeys with high cholesterol diets. Importantly, we found that a high cholesterol diet increases adverse histopathologic events and plaque burden, which are closely associated with changes in cholesterol, glucose, diastolic blood pressure, pulse wave velocity and could be predicted by the changes in contrast enhancement, luminal diameter, and wall thickness determined by MRI. Further studies indicate that HCD reprograms the aged arterial wall via further damage of endothelial integrity and macrophage infiltration, foam cell formation, and fat or calcium deposits, contributing to a vulnerable thickened intima for the accelerated occurrence of adverse histopathologic events, including plaque burden. Notably, comprehensive quantitative proteomic studies were designed to analyze proteomic changes of carotid arteries in the different conditions. We have found 8 proteins are less abundant and 12 proteins are more abundant in old animals. For example, the proteins which are less abundant are Protein S100-A6, Isocitrate dehydrogenase NADP, Myelin P0 protein, Myosin-7, Aldo-keto reductase family 1 member B10, Protein S100-A4, Calmodulin and SPATS2-like protein. The proteins which are more abundant include Programmed cell death protein 6, Periostin, Apolipoprotein E, Erythrocyte band 7 integral membrane protein, Translation initiation factor IF-2 Complement component C9, Lactadherin( MFG-E8), Apolipoprotein C-I Vitronectin, Annexin A7, Cysteine and glycine-rich protein 2, Serine protease HTRA1,Serum amyloid P-component and Complement C1q tumor necrosis factor-related protein. In addition, we have also identified protein abundance changes using 2-D DIGE. More than 20 protein spots have been found that have different abundance between protein extract of carotid arteries from young and old monkeys. Importantly, we have performed micro RNA arrays of carotid arteries in monkeys with or without high cholesterol diets. In brief, hybridization of Hy3-labeled RNA was performed on the miRCURY LNA miRNA microarray slide (version 11.0-other species), which contains 505 miRNA probes, including 473 monkey miRNA genes, spotted in quadruplicate. The microarray data normalized by Z score transformation were analyzed in the calculation of significant changes in gene expression changes between different samples and conditions. There are totally 91 modified miRs, via a continuous age analysis. 15 miRs significantly change with aging, 67 miRs are significantly different in abundance associated with high cholesterol diet treatment. Part of PCR validation results showed that miR-21, miR-34a, miR-155, miR-210, miR-199a, miR-199a-3p, miR-423-5p, let-7i are indeed modulated with age and/or high cholesterol diet treatment. Studies of bioroles of those miRs in aging and atherosclerosis are in progress. Furthermore, we found that carotid arteries of aged rhesus macaques exhibit significant oxidative stress (as indicated by the increased 8-iso-PGF2 and 4-HNE content and decreased glutathione and ascorbate levels) as compared with vessels of young macaques that is associated with activation of the redox-sensitive proinflammatory transcription factor, nuclear factor-kappaB. However, age-related oxidative stress does not activate Nrf2 and does not induce Nrf2 target genes (NQO1, GCLC, and HMOX1). In cultured vascular smooth muscle cells (VSMCs) derived from young M mulatta, treatment with H(2)O(2) and high glucose significantly increases transcriptional activity of Nrf2 and upregulates the expression of Nrf2 target genes. In contrast, in cultured vascular smooth muscle cells cells derived from aged macaques, H(2)O(2)- and high glucose-induced Nrf2 activity and Nrf2-driven gene expression are blunted. High glucose-induced H(2)O(2) production was significantly increased in aged vascular smooth muscle cells compared with that in vascular smooth muscle cells from young M mulatta. We also analyzed the cytokine secretion profile of primary vascular smooth muscle cells (VSMCs) derived from young and aged Macaca mulatta. Aged VSMCs cultured in the absence of systemic factors exhibited significantly increased secretion of interleukin-1, MCP-1, and tumor necrosis factor compared with young control cells. Secretion of interleukin-6 also tended to increase in aged VSMCs. This age-associated proinflammatory shift in the cellular secretory phenotype was associated with an increased mitochondrial O(2)(-) production and nuclear factor -light-chain-enhancer of activated B cells activation. Treatment of aged VSMCs with a physiologically relevant concentration of resveratrol (1 M) exerted significant anti-inflammatory effects, reversing aging-induced alterations in the cellular cytokine secretion profile and inhibiting nuclear factor -light-chain-enhancer of activated B cells. Resveratrol also attenuated mitochondrial O(2)(-) production and upregulated the transcriptional activity of Nrf2 in aged VSMCs. Thus, in non-human primates, cell-autonomous activation of nuclear factor -light-chain-enhancer of activated B cells and expression of an inflammatory secretome likely contribute to vascular inflammation in aging. Resveratrol treatment prevents the proinflammatory properties of the aged VSMC secretome, an effect that likely contributes to the demonstrated vasoprotective action of resveratrol in animal models of aging.
|Mattison, Julie A; Wang, Mingyi; Bernier, Michel et al. (2014) Resveratrol prevents high fat/sucrose diet-induced central arterial wall inflammation and stiffening in nonhuman primates. Cell Metab 20:183-90|