Arteriosclerosis, Thrombosis, and Vascular Biology最新文献

Dysfunctional endothelium is increasingly recognized as a mechanistic link between cardiovascular risk factors and dementia, including Alzheimer disease. BACE1 (β-site amyloid-β precursor protein-cleaving enzyme 1) is responsible for β-processing of APP (amyloid-β precursor protein), the first step in the production of Aβ (amyloid-β) peptides, major culprits in the pathogenesis of Alzheimer disease. Under pathological conditions, excessive activation of BACE1 exerts detrimental effects on endothelial function by Aβ-dependent and Aβ-independent mechanisms. High local concentration of Aβ in the brain blood vessels is responsible for the loss of key vascular protective functions of endothelial cells. More recent studies recognized significant contribution of Aβ-independent proteolytic activity of endothelial BACE1 to the pathogenesis of endothelial dysfunction. This review critically evaluates existing evidence supporting the concept that excessive activation of BACE1 expressed in the cerebrovascular endothelium impairs key homeostatic functions of the brain blood vessels. This concept has important therapeutic implications. Indeed, improved understanding of the mechanisms of endothelial dysfunction may help in efforts to develop new approaches to the protection and preservation of healthy cerebrovascular function.

Ceramides, a group of biologically active sphingolipids, have been described as the new cholesterol given strong evidence linking high plasma ceramide with endothelial damage, risk for early adverse cardiovascular events, and development of cardiometabolic disease. This relationship has sparked great interest in investigating therapeutic targets with the goal of suppressing ceramide formation. However, the growing data challenge this paradigm of ceramide as solely eliciting detrimental effects to the cardiovascular system. Studies show that ceramides are necessary for maintaining proper endothelial redox states, mechanosensation, and membrane integrity. Recent work in preclinical models and isolated human microvessels highlights that the loss of ceramide formation can in fact propagate vascular endothelial dysfunction. Here, we delve into these conflicting findings to evaluate how ceramide may be capable of exerting both beneficial and damaging effects within the vascular endothelium. We propose a unifying theory that while basal levels of ceramide in response to physiological stimuli are required for the production of vasoprotective metabolites such as S1P (sphingosine-1-phosphate), the chronic accumulation of ceramide can promote activation of pro-oxidative stress pathways in endothelial cells. Clinically, the evidence discussed here highlights the potential challenges associated with therapeutic suppression of ceramide formation as a means of reducing cardiovascular disease risk.

Background: Despite being in an oxygen-rich environment, endothelial cells (ECs) use anaerobic glycolysis (Warburg effect) as the primary metabolic pathway for cellular energy needs. PFKFB (6-phosphofructo-2-kinase/fructose-2,6-biphosphatase)-3 regulates a critical enzymatic checkpoint in glycolysis and has been shown to induce angiogenesis. This study builds on our efforts to determine the metabolic regulation of ischemic angiogenesis and perfusion recovery in the ischemic muscle.

Methods: Hypoxia serum starvation (HSS) was used as an in vitro peripheral artery disease (PAD) model, and hind limb ischemia by femoral artery ligation and resection was used as a preclinical PAD model.

Results: Despite increasing PFKFB3-dependent glycolysis, HSS significantly decreased the angiogenic capacity of ischemic ECs. Interestingly, inhibiting PFKFB3 significantly induced the angiogenic capacity of HSS-ECs. Since ischemia induced a significant in PFKFB3 levels in hind limb ischemia muscle versus nonischemic, we wanted to determine whether glucose bioavailability (rather than PFKFB3 expression) in the ischemic muscle is a limiting factor behind impaired angiogenesis. However, treating the ischemic muscle with intramuscular delivery of D-glucose or L-glucose (osmolar control) showed no significant differences in the perfusion recovery, indicating that glucose bioavailability is not a limiting factor to induce ischemic angiogenesis in experimental PAD. Unexpectedly, we found that shRNA-mediated PFKFB3 inhibition in the ischemic muscle resulted in an increased perfusion recovery and higher vascular density compared with control shRNA (consistent with the increased angiogenic capacity of PFKFB3 silenced HSS-ECs). Based on these data, we hypothesized that inhibiting HSS-induced PFKFB3 expression/levels in ischemic ECs activates alternative metabolic pathways that revascularize the ischemic muscle in experimental PAD. A comprehensive glucose metabolic gene qPCR arrays in PFKFB3 silenced HSS-ECs, and PFKFB3-knock-down ischemic muscle versus respective controls identified UGP2 (uridine diphosphate-glucose pyrophosphorylase 2), a regulator of protein glycosylation and glycogen synthesis, is induced upon PFKFB3 inhibition in vitro and in vivo. Antibody-mediated inhibition of UGP2 in the ischemic muscle significantly impaired perfusion recovery versus IgG control. Mechanistically, supplementing uridine diphosphate-glucose, a metabolite of UGP2 activity, significantly induced HSS-EC angiogenic capacity in vitro and enhanced perfusion recovery in vivo by increasing protein glycosylation (but not glycogen synthesis).

Conclusions: Our data present that inhibition of maladaptive PFKFB3-driven glycolysis in HSS-ECs is necessary to promote the UGP2-uridine diphosphate-glucose axis that enhances ischemic angiogenesis and perfusion recovery in experimental PAD.

Background: Vascular smooth muscle cells (VSMCs) are highly plastic. Vessel injury induces a phenotypic transformation from differentiated to dedifferentiated VSMCs, which involves reduced expression of contractile proteins and increased production of extracellular matrix and inflammatory cytokines. This transition plays an important role in several cardiovascular diseases such as atherosclerosis, hypertension, and aortic aneurysm. TGF-β (transforming growth factor-β) is critical for VSMC differentiation and to counterbalance the effect of dedifferentiating factors. However, the mechanisms controlling TGF-β activity and VSMC phenotypic regulation under in vivo conditions are poorly understood. The extracellular matrix protein TN-X (tenascin-X) has recently been shown to bind TGF-β and to prevent it from activating its receptor.

Methods: We studied the role of TN-X in VSMCs in various murine disease models using tamoxifen-inducible SMC-specific knockout and adeno-associated virus-mediated knockdown.

Results: In hypertensive and high-fat diet-fed mice, after carotid artery ligation as well as in human aneurysmal aortae, expression of Tnxb, the gene encoding TN-X, was increased in VSMCs. Mice with smooth muscle cell-specific loss of TN-X (SMC-Tnxb-KO) showed increased TGF-β signaling in VSMCs, as well as upregulated expression of VSMC differentiation marker genes during vascular remodeling compared with controls. SMC-specific TN-X deficiency decreased neointima formation after carotid artery ligation and reduced vessel wall thickening during Ang II (angiotensin II)-induced hypertension. SMC-Tnxb-KO mice lacking ApoE showed reduced atherosclerosis and Ang II-induced aneurysm formation under high-fat diet. Adeno-associated virus-mediated SMC-specific expression of short hairpin RNA against Tnxb showed similar beneficial effects. Treatment with an anti-TGF-β antibody or additional SMC-specific loss of the TGF-β receptor reverted the effects of SMC-specific TN-X deficiency.

Conclusions: In summary, TN-X critically regulates VSMC plasticity during vascular injury by inhibiting TGF-β signaling. Our data indicate that inhibition of vascular smooth muscle TN-X may represent a strategy to prevent and treat pathological vascular remodeling.

Background: Vascular calcification is associated with increased mortality in patients with cardiovascular disease. Secondary calciprotein particles are believed to play a causal role in the pathophysiology of vascular calcification. The maturation time (T₅₀) of calciprotein particles provides a measure of serum calcification propensity. We compared T₅₀ between patients with ST-segment-elevated myocardial infarction and control subjects and studied the association of T₅₀ with cardiovascular risk factors and outcome.

Methods: T₅₀ was measured by nephelometry in 347 patients from the GIPS-III trial (Metabolic Modulation With Metformin to Reduce Heart Failure After Acute Myocardial Infarction: Glycometabolic Intervention as Adjunct to Primary Coronary Intervention in ST Elevation Myocardial Infarction: a Randomized Controlled Trial) and in 254 matched general population controls from PREVEND (Prevention of Renal and Vascular End-Stage Disease). We also assessed the association between T₅₀ and left ventricular ejection fraction, as well as infarct size, the incidence of ischemia-driven reintervention during 5 years of follow-up, and serum nitrite as a marker of endothelial dysfunction.

Results: Patients with ST-segment-elevated myocardial infarction had a significantly lower T₅₀ (ie, higher serum calcification propensity) compared with controls (T₅₀: 289±63 versus 338±56 minutes; P<0.001). In patients with ST-segment-elevated myocardial infarction, lower T₅₀ was associated with female sex, lower systolic blood pressure, lower total cholesterol, lower LDL (low-density lipoprotein) cholesterol, lower triglycerides, and higher HDL (high-density lipoprotein) cholesterol but not with circulating nitrite or nitrate. Ischemia-driven reintervention was associated with higher LDL (P=0.03) and had a significant interaction term for T₅₀ and sex (P=0.005), indicating a correlation between ischemia-driven reintervention and T₅₀ above the median in men and below the median in women, between 150 days and 5 years of follow-up.

Conclusions: Serum calcification propensity is increased in patients with ST-segment-elevated myocardial infarction compared with the general population, and its contribution is more pronounced in women than in men. Its lack of/inverse association with nitrite and blood pressure confirms T₅₀ to be orthogonal to traditional cardiovascular disease risk factors. Lower T₅₀ was associated with a more favorable serum lipid profile, suggesting the involvement of divergent pathways of calcification stress and lipid stress in the pathophysiology of myocardial infarction.

Background: Atherosclerosis is driven by the infiltration of the arterial intima by diverse immune cells and smooth muscle cells (SMCs). CD8⁺ T cells promote lesion growth during atherosclerotic lesion development, but their role in advanced atherosclerosis is less clear. Here, we studied the role of CD8⁺ T cells and their effects on SMCs in established atherosclerosis.

Methods: CD8⁺ T cells were depleted in (SMC reporter) low-density lipoprotein receptor-deficient (Ldlr^-/-) mice with established atherosclerotic lesions. Atherosclerotic lesion formation was examined, and single-cell RNA sequencing of aortic SMCs and their progeny was performed. Additionally, coculture experiments with primary aortic SMCs and CD8⁺ T cells were conducted.

Results: Although we could not detect differences in atherosclerotic lesion size, an increased plaque SMC content was noted in mice after CD8⁺ T-cell depletion. Single-cell RNA sequencing of aortic lineage-traced SMCs revealed contractile SMCs and a modulated SMC cluster, expressing macrophage- and osteoblast-related genes. CD8⁺ T-cell depletion was associated with an increased contractile but decreased macrophage and osteoblast-like gene signature in this modulated aortic SMC cluster. Conversely, exposure of isolated aortic SMCs to activated CD8⁺ T cells decreased the expression of genes indicative of a contractile SMC phenotype and induced a macrophage and osteoblast-like cell state. Notably, CD8⁺ T cells triggered calcium deposits in SMCs under osteogenic conditions. Mechanistically, we identified transcription factors highly expressed in modulated SMCs, including Runx1, to be induced by CD8⁺ T cells in cultured SMCs in an IFNγ (interferon-γ)-dependent manner.

Conclusions: We here uncovered CD8⁺ T cells to control the SMC phenotype in atherosclerosis. CD8⁺ T cells promote SMC dedifferentiation and drive SMCs to adopt features of macrophage-like and osteoblast-like, procalcifying cell phenotypes. Given the critical role of SMCs in atherosclerotic plaque stability, CD8⁺ T cells could thus be explored as therapeutic target cells during lesion progression.

Background: The chromatin-remodeling enzymes BRG1 (brahma-related gene 1) and CHD4 (chromodomain helicase DNA-binding protein 4) independently regulate the transcription of genes critical for vascular development, but their coordinated impact on vessels in late-stage embryos has not been explored.

Methods: In this study, we genetically deleted endothelial Brg1 and Chd4 in mixed background mice (Brg1^fl/fl;Chd4^fl/fl;VE-Cadherin-Cre), and littermates that were negative for Cre recombinase were used as controls. Tissues were analyzed by immunostaining, immunoblot, and flow cytometry. Quantitative reverse transcription polymerase chain reaction was used to determine gene expression, and chromatin immunoprecipitation revealed gene targets of BRG1 and CHD4 in cultured endothelial cells.

Results: We found Brg1/Chd4 double mutants grew normally but died soon after birth with small and compact lungs. Despite having normal cellular composition, distal air sacs of the mutant lungs displayed diminished ECM (extracellular matrix) components and TGFβ (transforming growth factor-β) signaling, which typically promotes ECM synthesis. Transcripts for collagen- and elastin-related genes and the TGFβ ligand Tgfb1 were decreased in mutant lung endothelial cells, but genetic deletion of endothelial Tgfb1 failed to recapitulate the small lungs and ECM defects seen in Brg1/Chd4 mutants. We instead found several ECM genes to be direct targets of BRG1 and CHD4 in cultured endothelial cells.

Conclusions: Collectively, our data highlight essential roles for endothelial chromatin-remodeling enzymes in promoting ECM deposition in the distal lung tissue during the saccular stage of embryonic lung development.