Endothelial Nitric Oxide Synthase (eNOS) S1176 phosphorylation status governs atherosclerotic lesion formation

Objective: We have previously demonstrated the in vivo importance of the Akt-eNOS substratekinase relationship, as defective postnatal angiogenesis characteristic of global Akt1-null mice is rescued when bred to gain-of-function eNOS S1176D mutant mice. While multiple studies support the cardioprotective role of endothelial NO generation, the causal role of Akt1-dependent eNOS S1176 phosphorylation during atherosclerotic plaque formation is not yet clear. Approach & Results: We herein bred congenic loss-of-function eNOS S1176A and gain-of function eNOS S1176D mutant mice to the proatherogenic Akt1-/-; ApoE-/- double knockout mice to definitively test the importance of Akt-mediated eNOS S1176 phosphorylation during atherogenesis. We find that a single amino acid substitution at the eNOS S1176 phosphorylation site yields divergent effects on atherosclerotic plaque formation, as an eNOS phospho-mimic aspartate (D) substitution at S1176 leads to decreased indices of atherosclerosis, even when on a proatherogenic Akt1 global deletion background. Conversely, mice harboring an unphosphorylatable mutation to alanine (S1176A) result in increased lipid deposition and cellular apoptosis, phenocopying the physiological consequence of eNOS deletion and/or impaired enzyme function. Furthermore, gene expression analyses of whole aortas indicate a combinatorial detriment from NO deficiency and Western Diet challenge, as loss-of-function eNOS SA mice on a high-fat and high-cholesterol diet present a unique expression pattern indicative of augmented T-cell activity when compared to eNOS S1176D mice. Conclusions: By using genetic epistasis approaches, we conclusively demonstrate that Akt mediated eNOS S1176 phosphorylation and subsequent activation remains to be the most physiologically relevant method of NO production to promote cardioprotective effects.

The scent of atherosclerosis

Vascular macrophages sense an odorant to induce atherosclerotic plaque formation

Blood microflora of patients with atherosclerotic vascular lesions and microflora of atherosclerotic plaques of carotid arteries

Aim of the study was to assess the frequency of microflora detection in the blood of patients with atherosclerotic vascular lesions and middle-aged patients without clinical symptoms of atherosclerosis. Compare the nature of microflora isolated from blood and the range of microorganisms isolated from atherosclerotic plaques (AP) of patients with carotid arteries atherosclerosis. Material and methods. The hemocultures of 118 men and 33 women with atherosclerosis (mean age 55.6 years) as well as 10 blood samples of 3 men and 7 women formed into a control group (mean age 37 years) were examined. Test samples were cultivated for 6 months. Tissue platings of carotid arteries AP in 11 women and 24 men of the main group (mean age 58.0) were incubated for 2 months. Methods based on Student’s t-test and Mann-Whitney U-test were used for statistical analysis of the obtained results. Results. Propionibacterium acnes hemocultures were detected in 9.9 % of patients from the main and control groups. In blood Staphylococcus epidermidis and Stenotrophomonas maltophylia were detected in 0.7 % of samples. P. acnes and Staphylococcus spp. cultures were obtained from AP in 34.3 and 45.7 %, including both microorganisms in 8.6 % of samples. Conclusions. P. acnes cultures are found equally frequently in the blood of atherosclerotic patients and patients not yet diagnosed with atherosclerosis. This study confirmed the presence of the same-name microorganisms in blood and atherosclerotic plaques. In 5.7 % (2 of 35) it was possible to extract simultaneously a P. acnes culture from two loci (atherosclerotic tissue and blood) in specific individuals. Further detailed research is required to study the etiological significance of the microbial factor in the atherosclerotic plaque formation.

SYNTAX and Coronary Artery Calcium Score Predict Atherosclerotic Plaque Formation in the Ascending Aorta

Background: There is no study about the relationship between the complexity of coronary artery disease (SYNTAX SCORE; SS), and coronary artery calcium (CAC) score, accompanied with aortic calcium score (ACS) levels. The objective of this study was to investigate the relationship between the preoperative SS and CAC scores accompanying ACS in isolated CABG patients and their postoperative clinical results. Methods: This study included 130 consecutive CABG patients. The mean age of the patients was 62.3 ± 8.62 years (range: 47-84 years). SS was measured using coronary angiography by an experienced cardiologist. We investigated the ACS accompanied with CAC scores using a multidetector computed tomography (MDCT) in the same session, preoperatively. Measurements of the CAC score and ACS were measured by an experienced radiologist, who was unaware of the study in the same session. In order to investigate aortic wall pathology in patients with positive aortic calcification, we provided aortic tissue samples prior to the proximal anastomosis of bypass grafts using No:11 scalpel. Results: Overall median SS was 39 ± 7.2 (range: 15-41). CAC score was zero in 34 patients (26.1%). For the patients with a CAC score of zero, the median SS was 32 ± 9.4. There was no evidence of aortic calcification or plaque formation in 62 patients (47.6%). In these patients, the median SS was 35.6 ± 11.3. No significant difference was found when both groups were compared and for those patients with a calcific score of zero (P = .85). The median CAC score and ACS were 238 ± 122 AU (range: 0-1238 AU) and 112 ± 40 AU (range: 0-730 AU), respectively (P = .0033). For patients with a CAC score and ACS ≥400 AU, the mean SYNTAX score was ≥ 37. SS was correlated with CAC score (R:0.585; P < .0001). SYNTAX was correlated with ACS (R:0.557; P < .001). In multivariate analysis of SS (OR 1.053, 95% CI: 1.003–1.106, P = .039), gender (OR 0.189, 95% CI: 0.053–0.678, P = 0.011), age (OR 1.454, 95% CI: 1.256–1.632, P = .012), and diabetes mellitus (OR 0.341, 95% CI: 1.006–1.124, P = .014) were independent predictors for CAC score and aortic calcification. Conclusions: CAC score and ACS are strongly correlated with the complexity of coronary arteries in CABG patients. The total CAC score (≥ 400 AU) was independently associated with the degree of SS (>37). To prevent MACCE and mortality in CABG patients, we suggest the measurement of CAC score accompanied with ACS using MDCT as a non-invasive method. Highlight points: • Atherosclerotic plaque formation in aorta and coronary arteries are the main risk factors for stroke and infarction in CABG operations. •SYNTAX score value and aortic atherosclerosis levels are directly correlated. •SYNTAX score may predict the complications due to atherosclerosis during heart surgery.

Anticholesterolemic Activity of Three Vegetal Extracts (Artichoke, Caigua, and Fenugreek) and Their Unique Blend

Hepatic-related diseases, in particular hyperlipidemia and hypercholesterolemia, are a thorn on the side of the national health institutes around the globe. Indeed, liver lipid and cholesterol dysregulation could lead to atherosclerotic plaque formation and cardiovascular diseases. Currently, statin administration and monacolin K consumption are the main therapies proposed to counter this alarming connection, but relevant side effects are known. To overcome this issue, safe nutraceutical formulations and/or vegetal extracts, endowed with anticholesterolemic activity, could be instrumental in hypercholesterolemia prevention and treatment. In the present work, the anticholesterolemic efficacy of three vegetal extracts used in traditional medicine (artichoke, caigua, and fenugreek), their unique blend (ACFB), and the monacolin K-containing red yeast extract (RYR), was investigated with an in vitro approach based on hepatic cell line HepG2. The impact on cholesterol of the three extracts, their blend, and RYR were investigated by determining hepatocyte total and free cholesterol and bile acids biosynthesis. According to our results, the anticholesterolemic activity of the vegetal extracts was confirmed, and a novel choleretic activity of caigua extract was evidenced. ACFB showed to be safer than RYR while showing a similar effect on total and free cholesterol and bile acids synthesis compared to it. The anticholesterolemic activity of the blend was obtained with lower vegetal extract concentrations compared with the single vegetal extract, potentially indicating an additive effect between the extracts. In conclusion, the vegetal extracts and their blend, ACFB, are safe and are endowed with anticholesterolemic activity, potentially providing complementary therapies to the statin-based ones for hyperlipidemia and hypercholesterolemia-related complications.

Loss of soluble guanylyl cyclase in platelets contributes to atherosclerotic plaque formation and vascular inflammation

Aim: The role of platelets in atherosclerosis remains incompletely understood. Variants in genes encoding the soluble guanylyl cyclase (sGC) in platelets are associated with coronary artery disease (CAD) risk. Here we sought to investigate the contribution of platelet sGC to atherosclerosis and the therapeutic potential of targeting sGC in atherosclerosis. Methods and Results: We genetically deleted sGC in platelets of atherosclerosis-prone Ldlr-/- mice. By intravital fluorescence microscopy such Pf4-Cre+Gucy1b1flox/floxLdlr-/- mice displayed enhanced leukocyte adhesion to atherosclerotic plaques in comparison with their litter mates. Moreover, histological and flow cytometry analyses revealed more numerous inflammatory leukocytes and larger plaque sizes in aortic tissue of Ldlr-/- mice lacking sGC in platelets. In vitro, supernatant from activated platelets lacking sGC promoted leukocyte adhesion to endothelial cells (EC) via enhanced EC activation. Using cytokine profiling, we identified reduced angiopoietin-1 release by Pf4-Cre+Gucy1b1flox/flox and human GUCY1A1 risk allele carrier platelets to be responsible for enhanced activation of EC and subsequent leukocyte adhesion. Pharmacological sGC stimulation increased platelet angiopoietin-1 release in vitro and reduced recruitment of adoptively transferred leukocytes in Ldlr-/- mice fed a Western diet. Pharmacological sGC stimulation further reduced atherosclerotic plaque formation and vascular inflammation. Conclusion: Loss of sGC in platelets contributes to atherosclerotic plaque formation via reduced release of the soluble factor angiopoietin-1 and, subsequently, enhanced leukocyte recruitment. Pharmacological sGC stimulation might represent a novel therapeutic strategy to prevent and treat CAD.

C1q/TNF-Related Protein 9 Attenuates Atherosclerosis by Inhibiting Hyperglycemia-Induced Endothelial Cell Senescence Through the AMPKα/KLF4 Signaling Pathway

Hyperglycemia-induced endothelial cell senescence has been widely reported to be involved in the pathogenesis of type 2 diabetes mellitus‒accelerated atherosclerosis. Thus, understanding the underlying mechanisms and identifying potential therapeutic targets for endothelial cell senescence are valuable for attenuating atherosclerosis progression. C1q/tumor necrosis factor-related protein 9 (CTRP9), an emerging potential cardiokine, exerts a significant protective effect with respect to atherosclerosis, particularly in endothelial cells. However, the exact mechanism by which CTRP9 prevents endothelial cells from hyperglycemia-induced senescence remains unclear. This study aimed to investigate the effects of CTRP9 on hyperglycemia-induced endothelial cell senescence and atherosclerotic plaque formation in diabetic apolipoprotein E knockout (ApoE KO) mice. Human umbilical vein endothelial cells (HUVECs) were cultured in normal glucose (5.5 mM) and high glucose (40 mM) with or without recombinant human CTRP9 protein (3 μg/ml) for 48 h. Purified lentiviruses overexpressing CTRP9 (Lv-CTRP9) and control vectors containing green fluorescent protein (Lv-GFP) were injected via the tail vein into streptozotocin-induced diabetic ApoE KO mice. Results revealed that exposure of HUVECs to HG significantly increased the expression of Krüppel-like factor 4 (KLF4) and cyclin-dependent kinase inhibitor p21 (p21) and decreased that of telomerase reverse transcriptase (TERT). Treatment with recombinant human CTRP9 protein protected HUVECs from HG-induced premature senescence and dysfunction. CTRP9 promoted the phosphorylation of AMP-activated kinase (AMPK), attenuated the expression of KLF4 and p21 induced by HG, and increased the expression of TERT in HUVECs. Furthermore, in the background of AMPKα knockdown or KLF4 activation, the protective effects of CTRP9 were abolished. In-vivo experiments showed that the overexpression of CTRP9 inhibited vascular senescence and reduced atherosclerotic plaque formation in ApoE KO mice with diabetes. In conclusion, we demonstrate that KLF4 upregulation plays a crucial role in HG-induced endothelial senescence. This anti-atherosclerotic effect of CTRP9 may be partly attributed to the inhibition of HG-induced endothelial senescence through an AMPKα/KLF4-dependent mechanism, suggesting that CTRP9 could benefit further therapeutic approaches for type 2 diabetes mellitus‒accelerated atherosclerosis.

Statin induced myalgia on high intensity statin in patients with Acute Coronary Syndrome

Background High intensity statins are recommended in patients with acute coronary syndrome. Statins inhibit atherosclerotic plaque formation in the coronary arteries and reducing the burden of ischemic heart disease, therefore decreasing the morbidity and mortality. Muscle symptoms are most common adverse effect of statins. Hence, the aim of this study is to determine the statin induced myalgia by the statin myalgia clinical score. Purpose To monitor the Statin induced myalgia on high intensity statin in patients with Acute Coronary Syndrome Methods This was an prospective observational study comprised of 418 patients with acute coronary syndrome who were commenced on high intensity statins (Rosuvastatin 20–40mg & Atorvastatin 40–80). These patients were followed at 4 weeks, 8 weeks and 12 weeks subsequently and the clinical myalgia score (SAMS-CI) was calculated at each visit to determine the statin induced myalgia. SAMS-CI was categorized as unlikely (2–6), possible (7–8) and probable (9–11) Results From 418 patients, 327 were males and 91 were females. Mean age was 55.6±11.14. Only 19 (7.63±1.8) patients developed muscle symptoms on high intensity statins (Rosuvastatin 20 mg and Atorvastatin 40 mg) on SAMS-CI Score. 5 patients were unlikely to develop myalgia on SAMS-CI and continued with the same dosage without any new symptoms. 6 patients were possible on SAMS-CI, therefore the dosage of these patients were decreased to moderate intensity statin (Rosuvastatin 10mg, Atorvastatin 20 mg), their symptoms were resolved and continued with the moderate intensity statins. Furthermore, Statin was hold in 8 patients in the probable category for 4 weeks until the resolution of symptoms followed by moderate intensity statins. Conclusion Statin induced myalgia is more reported in old aged and female patients. Most of the patients can better tolerate the lower range of high intensity statins with the similar benefits and should be prescribed in every patient FUNDunding Acknowledgement Type of funding sources: None.

Redefining residual inflammatory risk after acute coronary syndrome

Over the last decades, inflammation proved to play a pivotal role in atherosclerotic plaque formation, progression and destabilization. Several studies showed that the patients presenting with acute coronary syndrome are at increased risk of adverse cardiovascular events at both short- and long-term follow-up. Results from different clinical trials highlighted that a residual inflammatory risk exist and targeting inflammation is a successful strategy in selected cases associated to an increased inflammatory burden. Recently, the optimization of intracoronary and multimodality imaging allowed to also assess the entity of local inflammation, thus encouraging the individuation of plaque characteristics that portend a higher risk of future cardiovascular events. In this short review, we aim to highlight the role of systemic and local inflammation in acute coronary syndromes, to provide a summarized overview of the possible medical strategies applicable in selected cases and to underline the diagnostic and prognostic potential of multimodality imaging.