Plaque Content Research Articles

Quantitative analysis of aortic Na[18F]F uptake in macrocalcifications and microcalcifications in PET/CT scans.

Currently, computed tomography (CT) is used for risk profiling of (asymptomatic) individuals by calculating coronary artery calcium scores. Although this score is a strong predictor of major adverse cardiovascular events, this method has limitations. Sodium [18F]fluoride (Na[18F]F) positron emission tomography (PET) has shown promise as an early marker for atherosclerotic progression. However, evidence on Na[18F]F as a marker for high-risk plaques is limited, particularly on its presentation in clinical PET/CT. Besides, the relationship between microcalcifications visualized by Na[18F]F PET and macrocalcifications detectable on CT is unknown. To establish a match/mismatch score in the aorta between macrocalcified plaque content on CT and microcalcification Na[18F]F PET uptake. Na[18F]F-PET/CT scans acquired in our centre in 2019-2020 were retrospectively collected. The aorta of each low-dose CT was manually segmented. Background measurements were placed in the superior vena cava. The vertebrae were automatically segmented using an open-source convolutional neural network, dilated with 10mm, and subtracted from the aortic mask. Per patient, calcium and Na[18F]F-hotspot masks were retrieved using an in-house developed algorithm. Three match/mismatch analyses were performed: a population analysis, a per slice analysis, and an overlap score. To generate a population image of calcium and Na[18F]F hotspot distribution, all aortic masks were aligned. Then, a heatmap of calcium HU and Na[18F]F-uptake on the surface was obtained by outward projection of HU and uptake values from the centerline. In each slice of the aortic wall of each patient, the calcium mass score and target-to-bloodpool ratios (TBR) were calculated within the calcium masks, in the aortic wall except the calcium masks, and in the aortic wall in slices without calcium. For the overlap score, three volumes were identified in the calcium and Na[18F]F masks: volume of PET (PET+/CT-), volume of CT (PET-/CT+), and overlapping volumes (PET+/CT+). A Spearman's correlation analysis with Bonferroni correction was performed on the population image, assessing the correlation between all HU and Na[18F]F vertex values. In the per slice analysis, a paired Wilcoxon signed-rank test was used to compare TBR values within each slice, while an ANOVA with post-hoc Kruskal-Wallis test was employed to compare TBR values between slices. p-values<0.05 were considered significant. In total, 186 Na[18F]F-PET/CT scans were included. A moderate positive exponential correlation was observed between total aortic calcium mass and total aortic TBR (r=0.68, p<0.001). A strong positive correlation (r=0.77, p<0.0001) was observed between CT values and Na[18F]F values on the population image. Significantly higher TBR values were found outside calcium masks than inside calcium masks (p<0.0001). TBR values in slices where no calcium was present, were significantly lower compared with outside calcium and inside calcium (both p<0.0001). On average, only 3.7% of the mask volumes were overlapping. Na[18F]F-uptake in the aorta behaves similarly to macrocalcification detectable on CT. Na[18F]F-uptake values are also moderately correlated to calcium mass scores (match). Higher uptake values were found just outside macrocalcification masks instead of inside the macrocalcification masks (mismatch). Also, only a small percentage of the Na[18F]F-uptake volumes overlapped with the calcium volumes (mismatch).

ADAMTS-7 Modulates Atherosclerotic Plaque Formation by Degradation of TIMP-1.

The ADAMTS7 locus was genome-wide significantly associated with coronary artery disease. Lack of the ECM (extracellular matrix) protease ADAMTS-7 (A disintegrin and metalloproteinase-7) was shown to reduce atherosclerotic plaque formation. Here, we sought to identify molecular mechanisms and downstream targets of ADAMTS-7 mediating the risk of atherosclerosis. Targets of ADAMTS-7 were identified by high-resolution mass spectrometry of atherosclerotic plaques from Apoe-/- and Apoe-/-Adamts7-/- mice. ECM proteins were identified using solubility profiling. Putative targets were validated using immunofluorescence, in vitro degradation assays, coimmunoprecipitation, and Förster resonance energy transfer-based protein-protein interaction assays. ADAMTS7 expression was measured in fibrous caps of human carotid artery plaques. In humans, ADAMTS7 expression was higher in caps of unstable as compared to stable carotid plaques. Compared to Apoe-/- mice, atherosclerotic aortas of Apoe-/- mice lacking Adamts-7 (Apoe-/-Adamts7-/-) contained higher protein levels of Timp-1 (tissue inhibitor of metalloprotease-1). In coimmunoprecipitation experiments, the catalytic domain of ADAMTS-7 bound to TIMP-1, which was degraded in the presence of ADAMTS-7 in vitro. ADAMTS-7 reduced the inhibitory capacity of TIMP-1 at its canonical target MMP-9 (matrix metalloprotease-9). As a downstream mechanism, we investigated collagen content in plaques of Apoe-/- and Apoe-/-Adamts7-/- mice after a Western diet. Picrosirius red staining of the aortic root revealed less collagen as a readout of higher MMP-9 activity in Apoe-/- as compared to Apoe-/- Adamts7-/- mice. To facilitate high-throughput screening for ADAMTS-7 inhibitors with the aim of decreasing TIMP-1 degradation, we designed a Förster resonance energy transfer-based assay targeting the ADAMTS-7 catalytic site. ADAMTS-7, which is induced in unstable atherosclerotic plaques, decreases TIMP-1 stability reducing its inhibitory effect on MMP-9, which is known to promote collagen degradation and is likewise associated with coronary artery disease. Disrupting the interaction of ADAMTS-7 and TIMP-1 might be a strategy to increase collagen content and plaque stability for the reduction of atherosclerosis-related events.

A novel dammarane triterpenoid alleviates atherosclerosis by activating the LXRα pathway

BackgroundWe have previously demonstrated that ginsenoside compound K can attenuate the formation of atherosclerotic lesions. Therefore, ginsenoside compound K has potential for atherosclerosis therapy. How to improve the druggability and enhance the antiatherosclerotic activity of ginsenoside compound K are the core problems in the prevention and treatment of atherosclerosis. CKN is a ginsenoside compound K derivative that was previously reported to have excellent antiatherosclerotic activity in vitro, and we have applied for international patents for it.MethodsMale C57BL/6 ApoE−/− mice were fed a high-fat and high-choline diet to induce atherosclerosis and were subjected to in vivo studies. In vitro, the CCK-8 method was applied to evaluate cytotoxicity in macrophages. Foam cells were utilized, and cellular lipid determination was performed for in vitro studies. The area of atherosclerotic plaque and fatty infiltration of the liver were measured by image analysis. Serum lipid and liver function were determined by a seralyzer. Immunofluorescence and western blot analysis were conducted to explore the alterations in the expression levels of lipid efflux-related proteins. Molecular docking, reporter gene experiments and cellular thermal shift assays were used to verify the interaction between CKN and LXRα.ResultsAfter confirming the therapeutic effects of CKN, molecular docking, reporter gene experiments and cellular thermal shift assays were used to predict and investigate the antiatherosclerotic mechanisms of CKN. CKN exhibited the greatest potency, with a 60.9% and 48.1% reduction in en face atherosclerotic lesions on the thoracic aorta and brachiocephalic trunk, reduced plasma lipid levels and decreased foam cell levels in the vascular plaque content in HHD-fed ApoE−/− mice. Moreover, CKN in the present study may exert its antiatherosclerotic effects through activated ABCA1 by promoting LXRα nuclear translocation and reducing the adverse effects of LXRα activation.ConclusionsOur results revealed that CKN prevented the formation of atherosclerosis in ApoE−/− mice by activating the LXRα pathway.

The association between circulating lipoprotein subfractions and lipid content in coronary atheromatous plaques assessed by near-infrared spectroscopy.

Lipid content in coronary atheromatous plaques, measured by near-infrared spectroscopy (NIRS), can predict the risk of future coronary events. Biomarkers that reflect lipid content in coronary plaques may therefore improve coronary artery disease (CAD) risk assessment. We aimed to investigate the association between circulating lipoprotein subfractions and lipid content in coronary atheromatous plaques in statin-treated patients with stable CAD undergoing percutaneous coronary intervention. 56 patients with stable CAD underwent three-vessel imaging with NIRS when feasible. The coronary artery segment with the highest lipid content, defined as themaximum lipid core burden index within any4mmlength across the entire lesion (maxLCBI4mm), was defined as target segment. Lipoprotein subfractions and Lipoprotein a (Lp(a)) were analyzed in fasting serum samples by nuclear magnetic resonance spectroscopy and by standard in-hospital procedures, respectively. Penalized linear regression analyses were used to identify the best predictors of maxLCBI4mm. The uncertainty of the lasso estimates was assessed as the percentage presence of a variable in resampled datasets by bootstrapping. Only modest evidence was found for an association between lipoprotein subfractions and maxLCBI4mm. The lipoprotein subfractions with strongest potential as predictors according to the percentage presence in resampled datasets were Lp(a) (78.1 % presence) and free cholesterol in the smallest high-density lipoprotein (HDL) subfractions (74.3 % presence). When including established cardiovascular disease (CVD) risk factors in the regression model, none of the lipoprotein subfractions were considered potential predictors of maxLCBI4mm. In this study, serum levels of Lp(a) and free cholesterol in the smallest HDL subfractions showed the strongest potential as predictors for lipid content in coronary atheromatous plaques. Although the evidence is modest, our study suggests that measurement of lipoprotein subfractions may provide additional information with respect to coronary plaque composition compared to traditional lipid measurements, but not in addition to established risk factors. Further and larger studies are needed to assess the potential of circulating lipoprotein subfractions as meaningful biomarkers both for lipid content in coronary atheromatous plaques and as CVD risk markers.

The association between circulating lipoprotein subfractions and lipid content in coronary atheromatous plaques assessed by near-infrared spectroscopy

Abstract Funding Acknowledgements Type of funding sources: Foundation. Main funding source(s): Norwegian Health Association The Liaison Committee for education, research and innovation in Central Norway. Background Lipid content in coronary atheromatous plaques, measured by near-infrared spectroscopy (NIRS), can predict the risk of future coronary events. Biomarkers that reflect lipid content in coronary plaques may therefore improve coronary artery disease (CAD) risk assessment, and lipoprotein subfractions could potentially add value beyond traditional lipid measurements. Purpose We aimed to investigate the association between circulating lipoprotein subfractions and lipid content in coronary atheromatous plaques in statin-treated patients with stable CAD undergoing percutaneous coronary intervention. Methods 56 patients with stable CAD underwent three vessel imaging with NIRS when feasible. The coronary artery segment with the highest lipid content, defined as the maximum lipid core burden index within any 4mm length across the entire lesion (maxLCBI4mm), was selected as target segment. Lipoprotein subfractions were analyzed in fasting serum samples by nuclear magnetic resonance spectroscopy. Penalized linear regression analyses (lasso) were used to identify the best predictors of maxLCBI4mm. The uncertainty of the lasso estimates was assessed as the percentage of the resampled datasets by bootstrapping. Results Only modest evidence was found for an association with lipoprotein subfractions and maxLCBI4mm. The lipoprotein subfractions with strongest potential as predictors according to the percentage presence in resampled datasets were lipoprotein a (Lp(a)) (reg.coeff = 57.0; 78.1 % presence) and free cholesterol in the smallest high-density lipoprotein (HDL) subfractions (reg.coeff = 36.5; 74.3 % presence). When including established cardiovascular risk factors in the regression model, none of the lipoprotein subfractions was considered potential predictors of maxLCBI4mm. Conclusion In this study, Lp(a) and free cholesterol in the smallest HDL subfractions in serum showed the strongest potential as predictors for lipid content in coronary atheromatous plaques. Although the evidence is modest, our study suggests that measurements lipoprotein subfractions may provide additional information with respect to coronary plaque composition compared to traditional lipid measurements, but not in addition to established risk factors. Future studies are needed to verify our findings, and to investigate the clinical implication of lipoprotein subfractions as meaningful biomarkers for lipid-rich plaques and enhanced risk prediction in CAD.

Abstract 313: Finasteride Delays Atherosclerosis Progression and Reduces Hepatic Inflammation in Low-density Lipoprotein Receptor-deficient Mice

Finasteride is commonly prescribed to treat benign hyperplasia and male-pattern baldness in cis men and, more recently, trans individuals. However, the effect of finasteride on cardiovascular disease remains elusive. We evaluated the role of finasteride on atherosclerosis using low-density lipoprotein receptor-deficient ( Ldlr -/- ) mice fed a Western diet containing increasing doses of finasteride for 12 weeks. We did not observe changes in food intake, body weight, or liver markers for toxicity for any of our experimental conditions. At the highest dose, finasteride reduced plasma cholesterol and triglycerides to approximately 25% compared to the control group. Plaque content at the aortic arch and aortic root level showed a reduction in lesion burden. Finasteride reduced monocytosis, monocyte recruitment to the lesion, macrophage lesion content, and necrotic area. RNA sequencing analysis revealed a downregulation of inflammatory pathways and an upregulation of bile acid metabolism, oxidative phosphorylation, and cholesterol pathways in the liver of mice taking finasteride. Our data unveil finasteride as a potential treatment to delay cardiovascular disease by its effects on plasma lipid profile.

ATP-binding cassette G1 membrane transporter-mediated cholesterol efflux capacity influences coronary atherosclerosis and cardiovascular risk in Rheumatoid Arthritis

ObjectivesCholesterol efflux capacity (CEC) measures the ability of high-density lipoprotein (HDL) to remove cholesterol from macrophages and reduce the lipid content of atherosclerotic plaques. CEC inversely associated with cardiovascular risk beyond HDL-cholesterol levels. CEC through the ATP-binding-cassette G1 (ABCG1) membrane transporter is impaired in rheumatoid arthritis (RA). We evaluated associations of ABCG1-CEC with coronary atherosclerosis, plaque progression and cardiovascular risk in RA. MethodsCoronary atherosclerosis (noncalcified, partially, fully-calcified, low-attenuation plaque) was assessed with computed tomography angiography in 140 patients and reevaluated in 99 after 6.9 ± 0.3 years. Cardiovascular events including acute coronary syndromes, stroke, cardiovascular death, claudication, revascularization and hospitalized heart failure were recorded. ABCG1-CEC was measured in Chinese hamster ovary cells as percentage of effluxed over total intracellular cholesterol. ResultsABCG1-CEC inversely associated with extensive atherosclerosis (≥5 plaques) (adjusted odds ratio 0.50 [95% CI 0.28–0.88]), numbers of partially-calcified (rate ratio [RR] 0.71 [0.53–0.94]) and low-attenuation plaques (RR 0.63 [0.43–0.91] per standard deviation increment). Higher ABCG1-CEC predicted fewer new partially-calcified plaques in patients with lower baseline and time-averaged CRP and fewer new noncalcified and calcified plaques in those receiving higher mean prednisone dose. ABCG1-CEC inversely associated with events in patients with but not without noncalcified plaques, with <median but not higher CRP and in prednisone users but not nonusers (p-for-interaction = 0.021, 0.033 and 0.008 respectively). ConclusionABCG1-CEC inversely associated with plaque burden and vulnerability, and plaque progression conditionally on cumulative inflammation and corticosteroid dose. ABCG1-CEC inversely associated with events specifically in patients with noncalcified plaques, lower inflammation and in prednisone users.

Myeloid-restricted CD68 deficiency attenuates atherosclerosis via inhibition of ROS-MAPK-apoptosis axis

In atherosclerosis, macrophages derived from blood monocytes contribute to non-resolving inflammation, which subsequently primes necrotic core formation, and ultimately triggers acute thrombotic vascular disease. Nevertheless, little is known about how inflammatory cells, especially the macrophages fuel atherosclerosis. CD68, a unique class D scavenger receptor (SRD) family member, is specifically expressed in monocytes/macrophages and remarkably up-regulated upon oxidized low-density lipoprotein (ox-LDL) stimulation. Nonetheless, whether and how myeloid-specific CD68 affects atherosclerosis remains to be defined. To determine the essential in vivo role and mechanism linking CD68 to atherosclerosis, we engineered global and myeloid-specific CD68-deficient mice on an ApoE-null background. On Western diet, both the mice with global and the myeloid-restricted deletion of CD68 on ApoE-null background attenuated atherosclerosis, accompanied by diminished immune/inflammatory cell burden and necrotic core content, but increased smooth muscle cell content in atherosclerotic plaques. In vitro experiments revealed that CD68 deficiency in macrophages resulted in attenuated ox-LDL-induced macrophage apoptosis. Additionally, CD68 deficiency suppressed ROS production, while removal of ROS can markedly reversed this effect. We further showed that CD68 deficiency affected apoptosis through inactivation of the mitogen-activated protein kinase (MAPK) pathway. Our findings establish CD68 as a macrophage lineage-specific regulator of “ROS-MAPK-apoptosis” axis, thus providing a previously unknown mechanism for the prominence of CD68 as a risk factor for coronary artery disease. Its therapeutic inhibition may provide a potent lever to alleviate the cardiovascular disease.

Re-Visiting Antioxidant Therapy in Murine Advanced Atherosclerosis with Brussels Chicory, a Typical Vegetable in Mediterranean Diets.

Brussels chicory, a typical vegetable in Mediterranean diets, has been recently reported to stabilize advanced atherosclerotic plaques in the brachiocephalic artery of apoE-deficient (Apoe-/-) mice. Herein, we investigated whether Brussels chicory can stabilize advanced plaques in the aorta via improving oxidative stress. Thirty week old Apoe-/- mice were fed the AIN-93G diet or supplemented with 0.5% freeze-dried Brussels chicory for twenty weeks. Aortic plaque size and stability, aortic relaxation, monocyte adhesion to aortic endothelium, free radicals, and enzymatic and non-enzymatic factors involved in free radical production and elimination in aorta and serum were measured. Brussels chicory consumption did not alter aortic plaque size, however, it stabilized aortic plaques, promoted aortic relaxation, and also inhibited monocyte adhesion to aortic endothelium. Moreover, this administration reduced oxidized LDL (ox-LDL) and 4-hydroxynonenal (4-HNE) content in aortic plaques, associated with inhibited aortic NADPH oxidase (NOX) and uncoupled endothelial nitric oxide synthase (eNOS)-mediated free radical production. However, Brussels chicory consumption did not appreciably alter aortic and serum superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) activities, aortic glutathione (GSH), as well as serum non-enzymatic antioxidants, such as bilirubin, uric acid, and GSH. Collectively, improved oxidative stress might contribute to the atheroprotective effect of Brussels chicory, supporting the prospect of the antioxidant therapy in advanced atherosclerosis progression.

MiR-320b is Associated with Lipid Content in Coronary Atheromatous Plaques

miR-320b is Associated with Lipid Content in Coronary Atheromatous Plaques

КОММЕМОРАТИВНЫЙ ДИСКУРС В КОНТЕКСТЕ ТРАНСФЕРА ЗНАНИЙ

The subject matter of the article is the content of one of the practices of commemorative discourse, namely memorial plaques put up in the 1920s - early 21st century. The article is aimed at analysing some mechanisms and means of knowledge transfer in commemorative discourse from an interdisciplinary perspective, using the example of memorial plaques. The work is based on the study of 1,500 memorial plaques in various cities of Russia, including national autonomies. Comparative analysis became the main research method. The ideas of cognitive approach made it possible to trace the processes of linguistic interpretation and understand the principles of information representation and knowledge transfer, which is relevant for the study of the informative and communicative function of commemorative discourse. The authors came to the following conclusions: the content of memorial plaques contributes to knowledge transfer; at that, this process is inseparable from the informative and communicative function of commemorative practices. The form of commemorative discourse under study can include all the main mechanisms of knowledge transfer; however, the focusing-defocusing mechanism is of primary importance, which is due to the peculiarities of a small-format text. All parts of the commemorative ensemble are involved in knowledge transfer. At the turn of the 21st century elements of the artistic design of memorial plaques, which expand and intensify verbal information, begin to play an increasingly significant role in this process. The results of the work can be used both in further theoretical research and in the practice of creating memorial plaques aimed at deepening their informative and communicative function.

Pyroglutamate-modified amyloid β(3-42) monomer has more β-sheet content than the amyloid β(1-42) monomer.

The aggregation of the amyloid β (Aβ) peptide is a major hallmark of Alzheimer's disease. This peptide can aggregate into oligomers, proto-fibrils and mature fibrils, which eventually assemble into amyloid plaques in vivo. Several post-translational modifications lead to the presence of different forms of the Aβ peptide in the amyloid plaques with different biophysical and biochemical properties. While the canonical forms Aβ(1-40) and Aβ(1-42) have been found to be the major components of amyloid plaques, N-terminally pyroglutamate-modified variants, specifically pE-Aβ(3-42), amount to a significant fraction of the total Aβ plaque content of AD brains. With increased hydrophobicity, these variants display a more pronounced aggregation behaviour in vitro which, together with their higher stability against degradation in vivo is thought to make them crucial molecular players in the aetiology of AD. The peptide monomers are the smallest assembly units, and play an important role in most of the individual molecular processes involved in amyloid fibril formation, such as primary and secondary nucleation and elongation. Understanding the monomeric conformational ensembles of the isoforms is important in unraveling observed differences in their bio-physico-chemical properties. Here we use enhanced and extensive molecular dynamics simulations to study the structural flexibility of the N-terminally truncated Pyroglutamate modified isomer of Aβ, pE-Aβ(3-42) monomer, and compared it with simulations of the Aβ(1-42) peptide monomer under the same conditions. We find significant differences, especially in the secondary structure and hydrophobic exposure, which might be responsible for their different behaviour in biophysical experiments.

Variation in aorta attenuation in contrast-enhanced CT and its implications for calcification thresholds.

CT contrast media improves vessel visualization but can also confound calcification measurements. We evaluated variance in aorta attenuation from varied contrast-enhancement scans, and quantified expected plaque detection errors when thresholding for calcification. We measured aorta attenuation (AoHU) in central vessel regions from 10K abdominal CT scans and report AoHU relationships to contrast phase (non-contrast, arterial, venous, delayed), demographic variables (age, sex, weight), body location, and scan slice thickness. We also report expected plaque segmentation false-negative errors (plaque pixels misidentified as non-plaque pixels) and false-positive errors (vessel pixels falsely identified as plaque), comparing a uniform thresholding approach and a dynamic approach based on local mean/SD aorta attenuation. Females had higher AoHU than males in contrast-enhanced scans by 65/22/20 HU for arterial/venous/delayed phases (p &lt; 0.001) but not in non-contrast scans (p &gt; 0.05). Weight was negatively correlated with AoHU by 2.3HU/10kg but other predictors explained only small portions of intra-cohort variance (R2 &lt; 0.1 in contrast-enhanced scans). Average AoHU differed by contrast phase, but considerable overlap was seen between distributions. Increasing uniform plaque thresholds from 130HU to 200HU/300HU/400HU produces respective false-negative plaque content losses of 35%/60%/75% from all scans with corresponding false-positive errors in arterial-phase scans of 95%/60%/15%. Dynamic segmentation at 3SD above mean AoHU reduces false-positive errors to 0.13% and false-negative errors to 8%, 25%, and 70% in delayed, venous, and arterial scans, respectively. CT contrast produces heterogeneous aortic enhancements not readily determined by demographic or scan protocol factors. Uniform CT thresholds for calcified plaques incur high rates of pixel classification errors in contrast-enhanced scans which can be minimized using dynamic thresholds based on local aorta attenuation. Care should be taken to address these errors and sex-based biases in baseline attenuation when designing automatic calcification detection algorithms intended for broad use in contrast-enhanced CTs.

Abstract 13040: Insulin-Like Growth Factor-1 Upregulates Larp6 via Translational and Post-Translational Mechanisms in Vascular Smooth Muscle Cells, Leading to Elevated Collagen Production

Introduction: Collagen content in atherosclerotic plaque is a hallmark of plaque stability. We have shown that insulin-like growth factor-1 (IGF1) increased collagen content in atherosclerotic plaques of Apoe -/- mice. We have identified LARP6 (La Ribonucleoprotein Domain Family, Member 6), a type I collagen mRNA-binding protein, as a mediator of IGF1-dependent type I collagen upregulation and increased mature fibril formation by smooth muscle cells (SMC), however specific mechanism has not been elucidated yet. Methods and Results: Micro-RNAs, including miR-1976 have been shown to bind to LARP6 mRNA. We found that SMC treatment with IGF1 significantly downregulated miR-1976 by 41±5% after 3h (P&amp;lt;0.05), consistent with the IGF1-induced LARP6 upregulation. In fact, miR-1976 mimic (20 nM) decreased LARP6 by 67% (P&amp;lt;0.05) and reduced type I procollagen protein levels by 85% (P&amp;lt;0.05). MiR-1976 mimic inhibits IGF1-induced LARP6 upregulation showing miR-1976 involvement in IGF1 effect on LARP6. It has been reported that LARP6-mediated promotion of procollagen synthesis may be regulated by phosphorylation of LARP6, therefore we investigated whether IGF1 induces LARP6 phosphorylation in cultured human aortic SMCs. For this experiment, SMCs were serum-starved and treated for 0-48h with 10 ng/mL IGF1. LARP6 protein was detected by immunoblots as a single 67kDa band in untreated SMC. However, after 6h of IGF1 treatment there was additional phosphorylated LARP6 band detected at 70 kDa. The levels of 70kDa LARP6 time-dependently increased up to 18h of treatment, with a concomitant increase of type I procollagen and collagen expression levels. Conclusions: In summary, our data suggests that (i) IGF1 downregulates miR-1976, thereby increasing LARP6 expression levels and (ii) IGF1 induces phosphorylation of LARP6, thereby promoting LARP6 activity. Our study revealed novel mechanisms underlying IGF1-induced type I collagen upregulation in vascular SMC and potentially in atherosclerotic plaques.

Abstract 9511: Sex Difference of Fibrous Cap Collagen Content in Patients With Coronary Artery Disease: An Intravascular Polarimetry Study

Introduction: Intracoronary optical coherence tomography (OCT) polarimetry (OCT-polarimetry) enables to provide polarization properties (birefringence and depolarization) of the tissues in the coronary arterial wall together with coronary plaque microstructures. Collagen and smooth muscle cells exhibit birefringence, while depolarization is increased by the presence of macrophages and lipid components. Hypothesis: We hypothesized that there exists sex difference in collagen content of coronary plaques by measuring polarization properties of the coronary arterial walls. Methods: We analyzed 41 coronary lesions imaged with intracoronary OCT-polarimetry in 38 patients with CAD (ACS, n = 24 and chronic CAD, n = 14). Conventional OCT analysis included lesion-level qualitative analysis (fibrous cap thickness, presence of plaque rupture, macrophages, thrombus, layered plaque, cholesterol crystals) and quantitative analysis (lipid-burden index, calcium arc and length, and vessel lumen area). Polarimetric analysis was performed on OCT cross-sectional images with the thinnest fibrous cap of the lesion, blinded to the conventional OCT analysis results. The measurements were compared between male (n = 27) and female (n =11) using a generalized linear model with generalized estimating equations. Results: In this study population, ACS was more frequently observed in female patients compared to male patients (91% v.s. 52%, p&lt;0.05). There was no significant difference in the conventional OCT measurements, while male patients exhibited a greater lipid-index compared to female patients. Although depolarization was comparable between the two groups, significantly lower birefringence was observed in female patients as compared to male patients (p-values &lt; 0.05). A GEE model demonstrated that birefringence was independently and positively associated with male sex (beta, 1.4; 95% confidence interval 0,42-2,4; p =0.005), even after adjusting for age, ACS, and lipid-burden index. Conclusions: This study is the first to report sex difference in the polarization properties of coronary plaque fibrous cap. Intracoronary OCT-polarimetry opens the possibility of elucidating the mechanism of plaque destabilization through collagen remodeling.

Abstract 14387: Early Atherosclerosis on CCTA: Insights From Artificial Intelligence-Enabled Quantitative CT Analysis of Serial Scans From the Paradigm Trial

Introduction: Non-obstructing small coronary plaques are not always appreciated during coronary CTA (CCTA) but may be identified by artificial intelligence-enabled quantitative CT (AI-QCT). It is unclear if these are true early atherosclerotic plaques or false positive findings. Hypothesis: Serial CCTA exams may provide insights into the natural history of these findings, as a plaque appearing in the same anatomical location and demonstrating interval change is likely real. Methods: 399 plaques measuring between 0.1 and 50.0 mm 3 were identified using AI-QCT in 83 patients with serial CCTA exams (N=166 exams) from the PARADIGM Trial. The total volume (TPV), calcified (CP), non-calcified (NCP) and low-density plaque (LD-NCP) volume, and % diameter stenosis of each plaque were recorded. Serial studies were processed in a blinded manner and linked after processing. The presence and change in plaque characteristics were assessed. Results: Overall 345/399 (86%) of small plaques were present in the same location on follow-up (mean interval 3.8 years), including 313/350 (89%) of plaques &gt;2.0mm 3 (Figure 1). Plaques were nonobstructive (&lt;=50%) in 99.5% of initial and 97.5% of follow-up studies. On follow-up, 82% (250/305) increased in size and 293/304 (96%) increased in calcified plaque content. Plaques in statin taking patients demonstrated higher median increase in CP (8.7 vs 4.0 mm 3 , p&lt;0.05) and transformed from all NCP to mixed NCP + CP at a higher rate (59.3 vs 43.9%, p=0.0027 Conclusions: Small coronary plaques depicted with serial AI-QCT are often present in the same location upon follow-up, increase in volume, and develop or increase calcification with statin therapy. While further data regarding the natural history and prognosis of these plaque is warranted, these findings provide a rationale for identifying early coronary atherosclerotic plaques.

Macrophage-specific deletion of neuropilin-2 inhibits inflammatory signalling and attenuates atherosclerotic plaque development in apolipoprotein e-deficient mice

Abstract Background Atherosclerosis is the underlying pathology in most cases of ischaemic heart disease, which itself remains the leading cause of mortality worldwide. Atherosclerosis is a multifaceted vascular disease characterised by arterial wall inflammation resulting from the subendothelial retention, and modification, of lipoproteins by macrophages. Macrophages exhibit a high degree of plasticity, and undergo polarisation to a pro-inflammatory phenotype in response to pro-inflammatory cytokines and other environmental stimuli, making them an attractive therapeutic target. Neuropilin-2 (NRP2) is a cell-surface co-receptor with essential roles in angiogenesis and axonal guidance, and is also expressed by macrophages. However, little is known about the role of NRP2 in macrophage function or atherosclerosis. Purpose To investigate whether NRP2 is important for macrophage involvement in atherosclerosis, we characterised the effects of conditional NRP2 deletion on macrophage function and atherosclerotic plaque development. Methods We generated mice with macrophage-specific (LysM-Cre) deletion of Nrp2, and lineage tracing via EYFP transgene expression, on the pro-atherogenic Apolipoprotein E-deficient background (Nrp2-KOMac, Apoe−/−, EYFP). These mice were fed a high-fat diet (HFD) for 16 weeks before their aortas were stained with Oil-red-O to assess plaque coverage. Plaque content was then characterised by sectioning and staining the aortic roots. The role of NRP2 in modulating pro-inflammatory macrophage polarisation and signalling, was assessed using functional assays and transcriptome analysis in Nrp2-KO bone marrow-derived macrophages (BMDMs). Results HFD-induced plaque development was significantly reduced in Nrp2-KOMac, Apoe−/−, EYFP mice, with 17.4% total aortic plaque coverage in Nrp2-KOMac, Apoe−/−, EYFP mice, compared to 24.9% in WTApoe−/−, EYFP mice (p=0.0021, n=10). Plaques from Nrp2-KOMac, Apoe−/−, EYFP mice also displayed features consistent with increased plaque stability, including reduced necrotic core area and plaque lipid content, and increased cap thickness. NRP2 was significantly upregulated upon M-CSF-mediated differentiation of bone marrow progenitors into BMDMs, and further upregulated by pro-inflammatory polarisation using LPS and IFN-gamma. Furthermore, upregulation of Il1β, Tnfα and Il6, induced by LPS and IFN-gamma, was significantly reduced in Nrp2-KO BMDMs, and MCP-1 induced migration was also decreased in these cells. Transcriptome analysis revealed that NF-kappaB signalling pathway genes, and genes regulating monocyte chemotaxis, were downregulated in Nrp2-KO BMDMs. Conclusion Macrophage-derived NRP2 is pro-atherogenic, likely resulting from its role in positively regulating pro-inflammatory signalling and macrophage migration. Targeting NRP2 expressed on the surface of macrophages could therefore offer a novel therapeutic approach for reducing the disease burden associated with atherosclerosis. Funding Acknowledgement Type of funding sources: Foundation. Main funding source(s): British Heart Foundation (MB PhD Studentship)

Interplanting of rice cultivars with high and low Cd accumulation can achieve the goal of “repairing while producing” in Cd-contaminated soil

Interplanting has been highlighted as a promising, cost-effective, and environmentally friendly solution for the remediation of contaminated soil. In this study, field experiments were conducted to study growth and cadmium (Cd) uptake in monoculture and interplanting systems with rice varieties Changliangyou 772 (C-772) and Changxianggu (Cho-ko-koku). And a pot culture experiment was conducted to investigate the response of the rhizosphere microecology of these two rice varieties. In the interplanting system of the field study, the Cd concentration in the grains of C-772 was significantly reduced (P < 0.05) from 0.30 mg kg−1 to 0.16 mg kg−1 and reached the national food safety standard of China (GB2762-2017, 0.20 mg kg−1), whereas the bioconcentration amount (BCA) per plant of Cd in Cho-ko-koku was significantly increased by 68.18 % (P < 0.05). The land equivalent ratio (LER) and Cd metal removal equivalent ratio (MRER) of the interplanting system were 1.03 and 1.05, illustrating that the interplanting system was superior in terms of yield and Cd elimination. In the pot experiment, the interplanting system significantly (P < 0.05) reduced the iron plaque content on the root surface and organic acids content in the rhizosphere environment of C-772 while markedly increasing those levels in Cho-ko-koku. At ripening stage, the interplanting system significantly decreased the rhizosphere available Cd concentration of C-772 from 0.38 mg kg−1 to 0.22 mg kg−1 (P < 0.05), while significantly increased the rhizosphere available Cd concentration of Cho-ko-koku from 0.27 mg kg−1 to 0.32 mg kg−1 (P < 0.05). Thus, Cd uptake of C-772 and Cho-ko-koku showed apparent differences. Oxalic and tartaric acid were identified as the most crucial factors affecting Cd uptake by C-772 and Cho-ko-koku in the interplanting system, respectively. In summary, this interplanting system is a promising planting pattern that can simultaneously improve land use efficiency and alleviate Cd pollution.

Abstract P1110: Identification Of Timp-1 As A Downstream Target Of The Coronary Artery Disease Risk Factor Adamts-7 In Atherosclerosis

Background: The gene encoding the extracellular matrix (ECM) protease ADAMTS-7 was associated with coronary artery disease (CAD) in genome-wide association studies. ADAMTS-7 is expressed at all stages in human plaques and mice lacking Adamts-7 displayed reduced atherosclerotic plaque formation. While these findings render ADAMTS-7 a promising therapeutic target, the underlying mechanisms remain unknown. Methods and Results: Here, we sought to identify downstream mechanisms of ADAMTS-7 in atherosclerotic plaque formation. Targets of Adamts-7 were identified by high-resolution mass spectrometry of atherosclerotic plaques in Apoe-/- and Apoe-/-Adamts7-/- mice. ECM proteins were identified using solubility profiling. The endogenous inhibitor of matrix metalloproteinases (MMP) Timp-1 was identified as a novel target of Adamts-7. Adamts-7 and Timp-1 were found to be co-localized in atherosclerotic plaques and co-immunoprecipitation (Co-IP) studies revealed TIMP-1 as the first putative target to bind to the catalytic domain of ADAMTS-7. In vitro degradation assays demonstrated that ADAMTS-7 degrades TIMP-1. Co-IP furthermore revealed less binding of TIMP-1 to its canonical target MMP-9 when ADAMTS-7 was present. In line, scaffolding and degradation of TIMP-1 by ADAMTS-7 impaired TIMP-1-mediated inhibition of MMP-9 in vitro. As a downstream mechanism, we investigated collagen content in atherosclerotic plaques of Apoe-/- and Apoe-/- Adamts7-/- mice after Western diet. Collagen stainings of the aortic root revealed less collagen as a readout of higher MMP-9 activity in Apoe-/- as compared to Apoe-/- Adamts7-/- mice. Since ADAMTS-7 might exert its pro-atherogenic effects through interaction with TIMP-1, we established a TIMP-1-ADAMTS-7-interaction assay based on Förster resonance energy transfer (FRET) to identify inhibitors of this protein-protein interaction. Conclusion and Outlook: TIMP-1 represents a novel downstream target of ADAMTS-7 that can explain the role of this novel risk factor in CAD. Our FRET-based protein-protein interaction assay may be used for high-throughput screening to identifiy inhibitors and prevent the initiation or slow the progression of atherosclerosis.

Real-Time Detection and Visualization of Amyloid-β Aggregates Induced by Hydrogen Peroxide in Cell and Mouse Models of Alzheimer's Disease.

Oxidative stress, caused by an imbalance between the production and the accumulation of reactive oxygen species (ROS), is a prominent cause of the neurotoxicity induced by aggregated amyloid-β (Aβ) in Alzheimer's disease (AD). Tools that can directly detect and monitor the presence and amount of Aβ-induced ROS are still lacking. We report herein the first Aβ-targeted ratiometric H2O2-responsive fluorescent probe for real-time detection and monitoring of the Aβ-induced H2O2 level in cell and AD mouse models. The H2O2-responsive probe is constructed based on a methylamino-substituted quinolinium-based cyanine as the fluorescence moiety and a phenylboronate ester as the sensing reaction site. This sensing probe exhibits a large emission wavelength shift of ∼87 nm upon reacting with H2O2, a high binding selectivity for Aβ, and a faster response toward H2O2 in the presence of Aβ, concomitant with an enhanced fluorescence intensity, hence greatly boosting the sensitivity of in-situ H2O2 detection. This biocompatible and nontoxic probe is capable of ratiometrically detecting and imaging endogenous H2O2 induced by Aβ in a neuronal cell model. Remarkably, this Aβ-targeted H2O2-responsive probe is also able to detect, monitor, and differentiate different Aβ-induced H2O2 levels in real time in different age groups of transgenic AD mice in which the cerebral H2O2 level increases age dependently concomitant with the plaque contents. Therefore, this smart probe can act as a powerful tool to diagnose high-risk subjects and diseased brains of AD and to further study the role of ROS in AD pathology.