Virginie Bolduc

Catechin treatment improves cerebrovascular flow-mediated dilation and learning abilities in atherosclerotic mice.

Severe dyslipidemia and the associated oxidative stress could accelerate the age-related decline in cerebrovascular endothelial function and cerebral blood flow (CBF), leading to neuronal loss and impaired learning abilities. We hypothesized that a chronic treatment with the polyphenol catechin would prevent endothelial dysfunction, maintain CBF responses, and protect learning abilities in atherosclerotic (ATX) mice. We treated ATX (C57Bl/6-LDLR(-/-)hApoB(+/+); 3 mo old) mice with catechin (30 mg · kg(-1) · day(-1)) for 3 mo, and C57Bl/6 [wild type (WT), 3 and 6 mo old] mice were used as controls. ACh- and flow-mediated dilations (FMD) were recorded in pressurized cerebral arteries. Basal CBF and increases in CBF induced by whisker stimulation were measured by optical coherence tomography and Doppler, respectively. Learning capacities were evaluated with the Morris water maze test. Compared with 6-mo-old WT mice, cerebral arteries from 6-mo-old ATX mice displayed a higher myogenic tone, lower responses to ACh and FMD, and were insensitive to NOS inhibition (P < 0.05), suggesting endothelial dysfunction. Basal and increases in CBF were lower in 6-mo-old ATX than WT mice (P < 0.05). A decline in the learning capabilities was also observed in ATX mice (P < 0.05). Catechin 1) reduced cerebral superoxide staining (P < 0.05) in ATX mice, 2) restored endothelial function by reducing myogenic tone, improving ACh- and FMD and restoring the sensitivity to nitric oxide synthase inhibition (P < 0.05), 3) increased the changes in CBF during stimulation but not basal CBF, and 4) prevented the decline in learning abilities (P < 0.05). In conclusion, catechin treatment of ATX mice prevents cerebrovascular dysfunctions and the associated decline in learning capacities.

Heart rate-associated mechanical stress impairs carotid but not cerebral artery compliance in dyslipidemic atherosclerotic mice

The cardiac cycle imposes a mechanical stress that dilates elastic carotid arteries, while shear stress largely contributes to the endothelium-dependent dilation of downstream cerebral arteries. In the presence of dyslipidemia, carotid arteries stiffen while the endothelial function declines. We reasoned that stiffening of carotid arteries would be prevented by reducing resting heart rate (HR), while improving the endothelial function would regulate cerebral artery compliance and function. Thus we treated or not 3-mo-old male atherosclerotic mice (ATX; LDLr(-/-):hApoB(+/+)) for 3 mo with the sinoatrial pacemaker current inhibitor ivabradine (IVA), the β-blocker metoprolol (METO), or subjected mice to voluntary physical training (PT). Arterial (carotid and cerebral artery) compliance and endothelium-dependent flow-mediated cerebral dilation were measured in isolated pressurized arteries. IVA and METO similarly reduced (P < 0.05) 24-h HR by ≈15%, while PT had no impact. As expected, carotid artery stiffness increased (P < 0.05) in ATX mice compared with wild-type mice, while cerebral artery stiffness decreased (P < 0.05); this paradoxical increase in cerebrovascular compliance was associated with endothelial dysfunction and an augmented metalloproteinase-9 (MMP-9) activity (P < 0.05), without changing the lipid composition of the wall. Reducing HR (IVA and METO) limited carotid artery stiffening, but plaque progression was prevented by IVA only. In contrast, IVA maintained and PT improved cerebral endothelial nitric oxide synthase-dependent flow-mediated dilation and wall compliance, and both interventions reduced MMP-9 activity (P < 0.05); METO worsened endothelial dysfunction and compliance and did not reduce MMP-9 activity. In conclusion, HR-dependent mechanical stress contributes to carotid artery wall stiffening in severely dyslipidemic mice while cerebrovascular compliance is mostly regulated by the endothelium.

Late chronic catechin antioxidant treatment is deleterious to the endothelial function in aging mice with established atherosclerosis.

Various antioxidants, including polyphenols, prevent the development of atherosclerosis in animal models, contrasting with the failure of antioxidants to provide benefits in patients with established atherosclerosis. We therefore tested in a mouse model the hypothesis that although catechin is atheroprotective in prevention, catechin brings no global vascular protection when initiated after established atherosclerosis, because aging associated with dyslipidemia has induced irreversible dysfunctions. To this end, LDLr(-/-); hApoB(+/+) atherosclerotic (ATX, 9 mo old) and pre-ATX (3 mo old) male mice were treated with catechin (30 mg x kg(-1) x day(-1)) up to 12 mo of age. Vascular function and endothelium/leukocyte interactions were studied at 12 mo old. The renal artery endothelium-dependent dilations were impaired with age whereas adhesion of leukocytes onto the native aortic endothelium was increased (P < 0.05). Aortic oxidative stress [reactive oxygen species (ROS)] increased (P < 0.05) at 3 mo in ATX and at 12 mo in wild-type mice. Aorta mRNA expression of NADPH oxidase increased, whereas that of manganese superoxide dismutase decreased in 12-mo-old ATX mice only. In mice with established ATX, catechin (from 9 to 12 mo) reduced (P < 0.05) by approximately 60% ROS without affecting plaque burden. Notably, catechin worsened endothelial dysfunction and further increased leukocyte adhesion (P < 0.05) in ATX mice. In contrast, the same catechin treatment reversed all age-related dysfunctions in wild-type mice. On the other hand, in pre-ATX mice treated for 9 mo with catechin, plaque burden was reduced by 64% (P < 0.05) and all vascular markers were normalized to the 3-mo-old values. These results demonstrate that an antioxidant treatment is deleterious in mice with established atherosclerosis.

Catechin prevents severe dyslipidemia-associated changes in wall biomechanics of cerebral arteries in LDLr−/−:hApoB+/+ mice and improves cerebral blood flow

Endothelial dysfunction and oxidative stress contribute to the atherosclerotic process that includes stiffening of large peripheral arteries. In contrast, our laboratory previously reported a paradoxical increase in cerebrovascular compliance in LDLr(-/-):hApoB(+/+) atherosclerotic (ATX) mice (7). We hypothesized that prevention of cerebral artery endothelial dysfunction with a chronic dietary antioxidant intake would normalize the changes in cerebral artery wall structure and biomechanics and prevent the decline in basal cerebral blood flow associated with atherosclerosis. Three-month-old ATX mice were treated, or not, for 3 mo with the polyphenol (+)-catechin (CAT; 30 mg·kg(-1)·day(-1)) and compared with wild-type controls. In isolated, pressurized cerebral arteries from ATX mice, CAT prevented endothelial dysfunction (deterioration of endothelium-dependent, flow-mediated dilations; P < 0.05), the inward hypertrophic structural remodeling (increase in the wall-to-lumen ratio; P < 0.05), and the rise in cerebrovascular compliance (rightward shift of the stress-strain curve measured in passive conditions, reflecting mechanical properties of the arterial wall; P < 0.05). Doppler optical coherence tomography imaging in vivo confirmed these findings, showing that cerebral compliance was higher in ATX mice and normalized by CAT (P < 0.05). CAT also prevented basal cerebral hypoperfusion in ATX mice (P < 0.05). Active remodeling of the cerebrovascular wall in ATX mice was further suggested by the increase (P < 0.05) in pro-metalloproteinase-9 activity, which was normalized by CAT. We conclude that, by preserving the endothelial function, a chronic treatment with CAT prevents the deleterious effect of severe dyslipidemia on cerebral artery wall structure and biomechanical properties, contributing to preserving resting cerebral blood flow.

Pulse pressure-dependent cerebrovascular eNOS regulation in mice.

Arterial blood pressure is oscillatory; whether pulse pressure (PP) regulates cerebral artery myogenic tone (MT) and endothelial function is currently unknown. To test the impact of PP on MT and dilation to flow (FMD) or to acetylcholine (Ach), isolated pressurized mouse posterior cerebral arteries were subjected to either static pressure (SP) or a physiological PP (amplitude: 30 mm Hg; frequency: 550 bpm). Under PP, MT was significantly higher than in SP conditions (p < 0.05) and was not affected by eNOS inhibition. In contrast, under SP, eNOS inhibition increased (p < 0.05) MT to levels observed under PP, suggesting that PP may inhibit eNOS. At a shear stress of 20 dyn/cm2, FMD was lower (p < 0.05) under SP than PP. Under SP, eNOS-dependent O 2 - / H 2 O 2 production contributed to FMD, while under PP, eNOS-dependent NO was responsible for FMD, indicating that PP favours eNOS coupling. Differences in FMD between pressure conditions were abolished after NOX2 inhibition. In contrast to FMD, Ach-induced dilations were higher (p < 0.05) under SP than PP. Reactive oxygen species scavenging reduced (p < 0.05) Ach-dependent dilations under SP, but increased (p < 0.05) them under PP; hence, under PP, Ach promotes ROS production and limits eNOS-derived NO activity. In conclusion, PP finely regulates eNOS, controlling cerebral artery reactivity.

Ivabradine and metoprolol differentially affect cardiac glucose metabolism despite similar heart rate reduction in a mouse model of dyslipidemia

While heart rate reduction (HRR) is a target for the management of patients with heart disease, contradictory results were reported using ivabradine, which selectively inhibits the pacemaker If current, vs. β-blockers like metoprolol. This study aimed at testing whether similar HRR with ivabradine vs. metoprolol differentially modulates cardiac energy substrate metabolism, a factor determinant for cardiac function, in a mouse model of dyslipidemia (hApoB+/+;LDLR-/-). Following a longitudinal study design, we used 3- and 6-mo-old mice, untreated or treated for 3 mo with ivabradine or metoprolol. Cardiac function was evaluated in vivo and ex vivo in working hearts perfused with 13C-labeled substrates to assess substrate fluxes through energy metabolic pathways. Compared with 3-mo-old, 6-mo-old dyslipidemic mice had similar cardiac hemodynamics in vivo but impaired (P < 0.001) contractile function (aortic flow: -45%; cardiac output: -34%; stroke volume: -35%) and glycolysis (-24%) ex vivo. Despite inducing a similar 10% HRR, ivabradine-treated hearts displayed significantly higher stroke volume values and glycolysis vs. their metoprolol-treated counterparts ex vivo, values for the ivabradine group being often not significantly different from 3-mo-old mice. Further analyses highlighted additional significant cardiac alterations with disease progression, namely in the total tissue level of proteins modified by O-linked N-acetylglucosamine (O-GlcNAc), whose formation is governed by glucose metabolism via the hexosamine biosynthetic pathway, which showed a similar pattern with ivabradine vs. metoprolol treatment. Collectively, our results emphasize the implication of alterations in cardiac glucose metabolism and signaling linked to disease progression in our mouse model. Despite similar HRR, ivabradine, but not metoprolol, preserved cardiac function and glucose metabolism during disease progression.

Synthetic reconstruction of dynamic blood flow in cortical arteries using optical coherence tomography for the evaluation of vessel compliance

Optical Coherence Tomography (OCT) has recently been used to produce 3D angiography of microvasculature in the rodent brain in-vivo and blood flow maps of large vessels. Key enabling developments were novel algorithms for detecting Doppler shifts produced by moving scatterers and new scanning protocols tailored to increase sensitivity to small flow speeds. These progresses were pushed by the need for a non invasive imaging modality to monitor quantitative blood flow at a higher resolution and a greater depth than could be achieved by other means. The rationale for this work originates from new hypotheses regarding the role of blood regulation in neurodegenerative diseases and from current investigations of animal models of vascular degeneration. In this work we demonstrate the synthetic reconstruction of dynamic blood flow in mice over the course of a single cardiac cycle in an 800μm wide by ~ 3mm deep B-Frame slice with a lateral resolution of 10μm and a depth resolution of 7μm. Images were taken using a cranial window over the exposed parietal bone of mice skull. Electrocardiography (ECG) recordings were co registered with the OCT A lines at high temporal resolution. QRS peak detection was then used to locate the time value of each A-line in the cardiac cycle and to reconstruct a synthetic temporal frame over one cardiac cycle. Doppler speed in this cardiac cycle was used to measure temporal variations of flow inside arteries and of their area. Three dimensional volume scans yielded measurements of quantitative blood flow on the same arteries. Using these informations a measure of compliance could be established. Comparing measures between atherosclerotic (ATX) and wild type (WT) mice revealed higher blood flow in WT mice, suggested lower systemic compliance in the ATX group but higher compliance of cerebral vasculature on these mice. These results are consistent with expectations showing that OCT is a potential tool for in-vivo arterial compliance evaluation.