The 9p21.3 coronary artery disease risk locus drives vascular smooth muscle cells to osteochondrogenic state.

Genome-wide association studies have identified common genetic variants at approximately 300 human genomic loci linked to CAD susceptibility. Among these genomic regions, the most impactful is the 9p21.3 CAD risk locus, which spans a 60 kb gene desert, encompasses about ~80 SNPs in high linkage disequilibrium, and has no defined function. We used induced pluripotent stem cell (iPSC) lines from risk and non-risk donors at 9p21.3, as well as isogenic lines with a full haplotype deletion. iPSC-derived vascular smooth muscle cells (iPSC-VSMCs) were used for single-cell transcriptomics. iPSC-VSMCs resemble the heterogeneity observed in human coronary arteries, establishing the robustness of this model. Our analysis revealed that the 9p21.3 risk haplotype prompts VSMCs to acquire a novel cellular state showing osteochondrogenic features, and we identified a set of signature genes crucial for defining this transcriptional program. Our study provides new insights into the 9p21.3 risk locus and its role in driving disease-prone states in VSMCs.

CD163+ Macrophages Induce Endothelial-to-Mesenchymal Transition in Atheroma.

BACKGROUND: Cell phenotype switching is increasingly being recognized in atherosclerosis. However, our understanding of the exact stimuli for such cellular transformations and their significance for human atherosclerosis is still evolving. Intraplaque hemorrhage is thought to be a major contributor to plaque progression in part by stimulating the influx of CD163+ macrophages. Here, we explored the hypothesis that CD163+ macrophages cause plaque progression through the induction of proapoptotic endothelial-to-mesenchymal transition (EndMT) within the fibrous cap. METHODS: Human coronary artery sections from CVPath's autopsy registry were selected for pathological analysis. Athero-prone ApoE-/- and ApoE-/-/CD163-/- mice were used for in vivo studies. Human peripheral blood mononuclear cell-induced macrophages and human aortic endothelial cells were used for in vitro experiments. RESULTS: In 107 lesions with acute coronary plaque rupture, 55% had pathological evidence of intraplaque hemorrhage in nonculprit vessels/lesions. Thinner fibrous cap, greater CD163+ macrophage accumulation, and a larger number of CD31/FSP-1 (fibroblast specific protein-1) double-positive cells and TUNEL (terminal deoxynucleotidyl transferase-dUTP nick end labeling) positive cells in the fibrous cap were observed in nonculprit intraplaque hemorrhage lesions, as well as in culprit rupture sections versus nonculprit fibroatheroma sections. Human aortic endothelial cells cultured with supernatants from hemoglobin/haptoglobin-exposed macrophages showed that increased mesenchymal marker proteins (transgelin and FSP-1) while endothelial markers (VE-cadherin and CD31) were reduced, suggesting EndMT induction. Activation of NF-κB (nuclear factor kappa ß) signaling by proinflammatory cytokines released from CD163+ macrophages directly regulated the expression of Snail, a critical transcription factor during EndMT induction. Western blot analysis for cleaved caspase-3 and microarray analysis of human aortic endothelial cells indicated that apoptosis was stimulated during CD163+ macrophage-induced EndMT. Additionally, CD163 deletion in athero-prone mice suggested that CD163 is required for EndMT and plaque progression. Using single-cell RNA sequencing from human carotid endarterectomy lesions, a population of EndMT was detected, which demonstrated significant upregulation of apoptosis-related genes. CONCLUSIONS: CD163+ macrophages provoke EndMT, which may promote plaque progression through fibrous cap thinning.

Adenosine kinase inhibition protects mice from abdominal aortic aneurysm via epigenetic modulation of VSMC inflammation.

AIM: Abdominal aortic aneurysm (AAA) is a common, serious vascular disease with no effective pharmacological treatment. The nucleoside adenosine plays an important role in modulating vascular homeostasis, which prompted us to determine whether adenosine kinase (ADK), an adenosine metabolizing enzyme, modulates AAA formation via control of intracellular adenosine level, and to investigate the underlying mechanisms. METHODS AND RESULTS: We used a combination of genetic and pharmacological approaches in murine models of AAA induced by calcium chloride (CaCl2) application or angiotensin II (Ang II) infusion to study the role of ADK in the development of AAA. In vitro functional assays were performed by knocking down ADK with adenovirus-short hairpin RNA in human vascular smooth muscle cells (VSMCs), and the molecular mechanisms underlying ADK function were investigated using RNA-sequencing, isotope tracing and chromatin immunoprecipitation quantitative polymerase chain reaction (ChIP-qPCR). Heterozygous deficiency of Adk protected mice from CaCl2- and Ang II-induced AAA formation. Moreover, specific knockout of Adk in VSMCs prevented Ang II-induced AAA formation, as evidenced by reduced aortic extracellular elastin fragmentation, neovascularization and aortic inflammation. Mechanistically, ADK knockdown in VSMCs markedly suppressed the expression of inflammatory genes associated with AAA formation, and these effects were independent of adenosine receptors. Metabolic flux and ChIP-qPCR results showed that ADK knockdown in VSMCs decreased S-adenosylmethionine (SAM)-dependent transmethylation, thereby reducing H3K4me3 binding to the promoter regions of the genes that are associated with inflammation, angiogenesis and extracellular elastin fragmentation. Furthermore, the ADK inhibitor ABT702 protected mice from CaCl2-induced aortic inflammation, extracellular elastin fragmentation and AAA formation. CONCLUSION: Our findings reveal a novel role for ADK inhibition in attenuating AAA via epigenetic modulation of key inflammatory genes linked to AAA pathogenesis.

Boosting treatment outcomes via the patient-practitioner relationship, treatment-beliefs or therapeutic setting. A systematic review with meta-analysis of contextual effects in chronic musculoskeletal pain.

OBJECTIVE: To ascertain whether manipulating contextual effects (e.g. interaction with patients, or beliefs about treatments) boosted the outcomes of non-pharmacological and non-surgicaltreatments for chronic primary musculoskeletal pain. DESIGN: Systematic review of randomized controlled trials. DATA SOURCES: We searched for trials in six databases, citation tracking, and clinical trials registers. We included trials that compared treatments with enhanced contextual effects with the same treatments without enhancement in adults with chronic primary musculoskeletal pain. DATA SYNTHESIS: The outcomes of interest were pain intensity, physical functioning, global ratings of improvement, quality of life, depression, anxiety, and sleep. We evaluated risk of bias and certainty of the evidence using Cochrane Risk of Bias tool 2.0 and the GRADE approach, respectively. RESULTS: Of 17637 records, we included 10 trials with 990 participants and identified 5 ongoing trials. The treatments were acupuncture, education, exercise training, and physical therapy. The contextual effects that were improved in the enhanced treatments were patient-practitioner relationship, patient beliefs and characteristics, therapeutic setting/environment, and treatment characteristics. Our analysis showed that improving contextual effects in non-pharmacological and non-surgical treatments may not make much difference on pain intensity (mean difference [MD] : -1.77, 95%-CI: [-8.71; 5.16], k = 7 trials, N = 719 participants, Scale: 0-100, GRADE: Low)) or physical functioning (MD: -0.27, 95%-CI: [-1.02; 0.49], 95%-PI: [-2.04; 1.51], k = 6 , N = 567, Scale: 0-10, GRADE: Low) in the short-term and at later follow-ups. Sensitivity analyses revealed similar findings. CONCLUSION: Whilst evidence gaps exist, per current evidence it may not be possible to achieve meaningful benefit for patients with chronic musculoskeletal pain by manipulating the context of non-pharmacological and non-surgical treatments. TRIAL REGISTRATION: This systematic review was prospectively registered in PROSPERO (registration number: CRD42023391601).

Single-Cell Gene-Regulatory Networks of Advanced Symptomatic Atherosclerosis.

BACKGROUND: While our understanding of the single-cell gene expression patterns underlying the transformation of vascular cell types during the progression of atherosclerosis is rapidly improving, the clinical and pathophysiological relevance of these changes remains poorly understood. METHODS: Single-cell RNA sequencing data generated with SmartSeq2 (≈8000 genes/cell) in 16 588 single cells isolated during atherosclerosis progression in Ldlr-/-Apob100/100 mice with human-like plasma lipoproteins and from humans with asymptomatic and symptomatic carotid plaques was clustered into multiple subtypes. For clinical and pathophysiological context, the advanced-stage and symptomatic subtype clusters were integrated with 135 tissue-specific (atherosclerotic aortic wall, mammary artery, liver, skeletal muscle, and visceral and subcutaneous, fat) gene-regulatory networks (GRNs) inferred from 600 coronary artery disease patients in the STARNET (Stockholm-Tartu Atherosclerosis Reverse Network Engineering Task) study. RESULTS: Advanced stages of atherosclerosis progression and symptomatic carotid plaques were largely characterized by 3 smooth muscle cells (SMCs), and 3 macrophage subtype clusters with extracellular matrix organization/osteogenic (SMC), and M1-type proinflammatory/Trem2-high lipid-associated (macrophage) phenotypes. Integrative analysis of these 6 clusters with STARNET revealed significant enrichments of 3 arterial wall GRNs: GRN33 (macrophage), GRN39 (SMC), and GRN122 (macrophage) with major contributions to coronary artery disease heritability and strong associations with clinical scores of coronary atherosclerosis severity. The presence and pathophysiological relevance of GRN39 were verified in 5 independent RNAseq data sets obtained from the human coronary and aortic artery, and primary SMCs and by targeting its top-key drivers, FRZB and ALCAM in cultured human coronary artery SMCs. CONCLUSIONS: By identifying and integrating the most gene-rich single-cell subclusters of atherosclerosis to date with a coronary artery disease framework of GRNs, GRN39 was identified and independently validated as being critical for the transformation of contractile SMCs into an osteogenic phenotype promoting advanced, symptomatic atherosclerosis.

A Large-Scale Genome-Wide Study of Gene-Sleep Duration Interactions for Blood Pressure in 811,405 Individuals from Diverse Populations.

Although both short and long sleep duration are associated with elevated hypertension risk, our understanding of their interplay with biological pathways governing blood pressure remains limited. To address this, we carried out genome-wide cross-population gene-by-short-sleep and long-sleep duration interaction analyses for three blood pressure traits (systolic, diastolic, and pulse pressure) in 811,405 individuals from diverse population groups. We discover 22 novel gene-sleep duration interaction loci for blood pressure, mapped to genes involved in neurological, thyroidal, bone metabolism, and hematopoietic pathways. Non-overlap between short sleep (12) and long sleep (10) interactions underscores the plausibility of distinct influences of both sleep duration extremes in cardiovascular health. With several of our loci reflecting specificity towards population background or sex, our discovery sheds light on the importance of embracing granularity when addressing heterogeneity entangled in gene-environment interactions, and in therapeutic design approaches for blood pressure management.

Association between BMD and coronary artery calcification: an observational and Mendelian randomization study.

Observational studies have reported inconsistent associations between bone mineral density (BMD) and coronary artery calcification (CAC). We examined the observational association of BMD with CAC in 2 large population-based studies and evaluated the evidence for a potential causal relation between BMD and CAC using polygenic risk scores (PRS), 1- and 2-sample Mendelian randomization (MR) approaches. Our study populations comprised 1414 individuals (mean age 69.9 yr, 52.0% women) from the Rotterdam Study and 2233 individuals (mean age 56.5 yr, 50.9% women) from the Framingham Heart Study with complete information on CAC and BMD measurements at the total body (TB-), lumbar spine (LS-), and femoral neck (FN-). We used linear regression models to evaluate the observational association between BMD and CAC. Subsequently, we compared the mean CAC across PRSBMD quintile groups at different skeletal sites. In addition, we used the 2-stage least squares regression and the inverse variance weighted (IVW) model as primary methods for 1- and 2-sample MR to test evidence for a potentially causal association. We did not observe robust associations between measured BMD levels and CAC. These results were consistent with a uniform random distribution of mean CAC across PRSBMD quintile groups (P-value > .05). Moreover, neither 1- nor 2-sample MR supported the possible causal association between BMD and CAC. Our results do not support the contention that lower BMD is (causally) associated with an increased CAC risk. These findings suggest that previously reported epidemiological associations of BMD with CAC are likely explained by unmeasured confounders or shared etiology, rather than by causal pathways underlying both osteoporosis and vascular calcification processes.

Decreased bone mineral density, the determinant of osteoporosis, and increased coronary artery calcification are common in people at an advanced age and share some common risk factors. Some studies have reported a higher risk for coronary artery calcification in people with osteoporosis than in people without, whereas others failed to find evidence for this relationship. Recently, Mendelian randomization has emerged as an important epidemiological tool that offers a simple way to distinguish causation, minimizing the confounding present in observational studies, leveraging individual genetic data and the findings from robust genome-wide association studies. We combined data from the participants of both the Rotterdam Study and the Framingham Heart Study, and did not observe sufficient evidence for the association between bone mineral density at different skeletal sites and coronary artery calcification. Also, when using Mendelian randomization, we concluded there was no causal relation between bone deterioration and the build-up of calcium in the coronary arteries. Although more research is needed, we conclude that the associations between decreased bone mineral density and increased coronary artery calcification reported in previous studies are likely attributed to other confounders rather than a causal relationship between these traits.

Exercise prescription variables predict reductions in pain intensity in adults with chronic low back pain: secondary analysis of a randomised controlled trial.

Objectives: The relationship between exercise training variables and clinical outcomes in low back pain (LBP) is unclear. The current study aimed to explore the relationship between exercise training parameters and pain intensity in individuals with chronic LBP. Methods: This study is a secondary analysis of a previously reported randomised controlled trial comparing the effects of general strength and conditioning to motor control exercises and manual therapy. This secondary analysis includes adults with chronic LBP (n=20) randomised to the general strength and conditioning programme only. Primary outcomes of this analysis were exercise training parameters (time under tension, rating of perceived exertion (RPE), session duration, session-RPE and training frequency) and pain intensity (0-100 mm visual analogue acale) measured every 2 weeks from baseline to 6 months follow-up. Linear mixed models with random effects (participants) and allowance for heterogeneity of variance (study date) were used to determine the association between pain intensity and training parameters over time. Results: Mean (95% CI) pain intensity decreased over time from baseline to 6 months follow-up by 10.7 (2.8 to 18.7) points (p=0.008). Over the 6-month intervention, lower pain intensity was associated with higher RPE (ß (95% CI) -27.168 (-44.265 to -10.071), p=0.002), greater time under tension (-0.029 (-0.056 to -0.001), p=0.040) and shorter session duration (1.938 (0.011 to 3.865), p=0.049). Conclusion: During 6 months of general strength and conditioning, lower pain intensity was associated with higher participant-reported training intensity, greater volume and shorter session duration. To ensure positive outcomes to exercise training, these variables should be monitored on a short-term basis. Trial registration number: ACTRN12615001270505.

Multi-ancestry genetic analysis of gene regulation in coronary arteries prioritizes disease risk loci.

Genome-wide association studies (GWASs) have identified hundreds of risk loci for coronary artery disease (CAD). However, non-European populations are underrepresented in GWASs, and the causal gene-regulatory mechanisms of these risk loci during atherosclerosis remain unclear. We incorporated local ancestry and haplotypes to identify quantitative trait loci for expression (eQTLs) and splicing (sQTLs) in coronary arteries from 138 ancestrally diverse Americans. Of 2,132 eQTL-associated genes (eGenes), 47% were previously unreported in coronary artery; 19% exhibited cell-type-specific expression. Colocalization revealed subgroups of eGenes unique to CAD and blood pressure GWAS. Fine-mapping highlighted additional eGenes, including TBX20 and IL5. We also identified sQTLs for 1,690 genes, among which TOR1AIP1 and ULK3 sQTLs demonstrated the importance of evaluating splicing to accurately identify disease-relevant isoform expression. Our work provides a patient-derived coronary artery eQTL resource and exemplifies the need for diverse study populations and multifaceted approaches to characterize gene regulation in disease processes.

The Impact of Exercise Prescription Variables on Intervention Outcomes in Musculoskeletal Pain: An Umbrella Review of Systematic Reviews.

BACKGROUND: Musculoskeletal pain conditions are the largest contributors to disability and healthcare burden globally. Exercise interventions improve physical function and quality of life in individuals with musculoskeletal pain, yet optimal exercise prescription variables (e.g. duration, frequency, intensity) are unclear. OBJECTIVE: We aimed to examine evidence gaps, methodological quality and exercise prescription recommendations in systematic reviews of exercise for musculoskeletal pain. METHODS: In our prospectively registered umbrella review, PubMed, SPORTDiscus, Cochrane Database of Systematic Reviews, EMBASE, and CINAHL were searched from inception to 14 February 2023. Backward citation tracking was performed. We included peer-reviewed, English language, systematic reviews and meta-analyses of randomized controlled trials (RCTs) and controlled clinical trials (CCTs) that compared exercise with conservative treatment, placebo or other exercise interventions in adults with musculoskeletal pain. Data were extracted from the following groups of reviews based on their reporting of exercise prescription data and analysis of the relationship between prescription variables and outcomes: (1) those that did not report any exercise prescription data, (2) those that reported exercise prescription data but did not perform a quantitative analysis and (3) those that performed a quantitative analysis of the relationship between exercise prescription variables and outcomes. Outcome measures were physical function, pain, mental health, adverse effects and adherence to treatment. AMSTAR-2 (A MeaSurement Tool to Assess systematic Reviews) was used to assess methodological quality. RESULTS: From 6757 records, 274 systematic reviews were included. 6.6% of reviews did not report any exercise prescription data, and only 10.9% quantitatively analyzed the relationship between prescription variables and the outcome(s). The overall methodological quality was critically low in 85% of reviews. CONCLUSION: High methodological quality evidence is lacking for optimal exercise training prescription variables in individuals with musculoskeletal pain. To better inform practice and evidence gaps, future systematic reviews should (1) identify optimum exercise prescription variables, for example, via dose-response (network) meta-analysis, (2) perform high-quality reviews per AMSTAR-2 criteria and (3) include outcomes of mental health, adverse events and exercise adherence. PROSPERO REGISTRATION NUMBER: CRD42021287440 ( https://www.crd.york.ac.uk/prospero/display_record.php?ID=CRD42021287440 ).

How to Interpret Effect Sizes for Biopsychosocial Outcomes and Implications for Current Research.

Biopsychosocial factors are associated with pain, but they can be difficult to compare. One way of comparing them is to use standardized mean differences. Previously, these effects sizes have been termed as small, medium, or large, if they are bigger than or equal to, respectively, .2, .5, or .8. These cut-offs are arbitrary and recent evidence showed that they need to be reconsidered. We argue it is necessary to determine cut-offs for each biopsychosocial factor. To achieve this, we propose 3 potential approaches: 1) examining, for each factor, how the effect size differs depending upon disease severity; 2) using an existing minimum clinically important difference to anchor the large effect size; and 3) define cut-offs by comparing data from people with and without pain. This is important for pain research, as exploring these methodologies has potential to improve comparability of biopsychosocial factors and lead to more directed treatments. We note assumptions and limitations of these methods that should also be considered. PERSPECTIVE: Standardized mean differences can estimate effect sizes between groups and could theoretically allow for comparison of biopsychosocial factors. However, common thresholds to define effect sizes are arbitrary and likely differ based on outcome. We propose methods that could overcome this and be used to derive biopsychosocial outcome-specific effect sizes.

Polygenic Risk Score Associates With Atherosclerotic Plaque Characteristics at Autopsy.

BACKGROUND: Polygenic risk scores (PRSs) for coronary artery disease (CAD) potentially improve cardiovascular risk prediction. However, their relationship with histopathologic features of CAD has never been examined systematically. METHODS: From 4327 subjects referred to CVPath by the State of Maryland Office Chief Medical Examiner for sudden death between 1994 and 2015, 2455 cases were randomly selected for genotyping. We generated PRS from 291 known CAD risk loci. Detailed histopathologic examination of the coronary arteries was performed in all subjects. The primary study outcome measurements were histopathologic plaque features determining severity of atherosclerosis, including %stenosis, calcification, thin-cap fibroatheromas, and thrombotic CAD. RESULTS: After exclusion of cases with insufficient DNA sample quality or with missing data, 954 cases (mean age, 48.8±14.7 years; 75.7% men) remained in the final study cohort. Subjects in the highest PRS quintile exhibited more severe atherosclerosis compared with subjects in the lowest quintile, with greater %stenosis (80.3%±27.0% versus 50.4%±38.7%; adjusted P<0.001) and a higher frequency of calcification (69.6% versus 35.8%; adjusted P=0.004) and thin-cap fibroatheroma (26.7% versus 9.5%; adjusted P=0.007). Even after adjustment for traditional CAD risk factors, subjects within the highest PRS quintile had higher odds of severe atherosclerosis (ie, ≥75% stenosis; adjusted odds ratio, 3.77 [95% CI, 2.10-6.78]; P<0.001) and plaque rupture (adjusted odds ratio, 4.05 [95% CI, 2.26-7.24]; P<0.001). Moreover, subjects within the highest quintile had higher odds of CAD-associated cause of death, especially among those aged ≤50 years (adjusted odds ratio, 4.08 [95% CI, 2.01-8.30]; P<0.001). No statistically significant associations were observed with plaque erosion after adjusting for covariates. CONCLUSIONS: This is the first autopsy study investigating associations between PRS and atherosclerosis severity at the histopathologic level in subjects with sudden death. Our pathological analysis suggests PRS correlates with plaque burden and features of advanced atherosclerosis and may be useful as a method for CAD risk stratification, especially in younger subjects.

The importance of context (placebo effects) in conservative interventions for musculoskeletal pain: A systematic review and meta-analysis of randomized controlled trials.

BACKGROUND AND OBJECTIVE: Contextual effects (e.g. patient expectations) may play a role in treatment effectiveness. This study aimed to estimate the magnitude of contextual effects for conservative, non-pharmacological interventions for musculoskeletal pain conditions. A systematic review and meta-analysis of randomized controlled trials (RCTs) that compared placebo conservative non-pharmacological interventions to no treatment for musculoskeletal pain. The outcomes assessed included pain intensity, physical functioning, health-related quality of life, global rating of change, depression, anxiety and sleep at immediate, short-, medium- and/or long-term follow-up. DATABASES AND DATA TREATMENT: MEDLINE, EMBASE, CINAHL, Web of Science Core Collection, CENTRAL and SPORTDiscus were searched from inception to September 2021. Trial registry searches, backward and forward citation tracking and searches for prior systematic reviews were completed. The Cochrane risk of bias 2 tool was implemented. RESULTS: The study included 64 RCTs (N = 4314) out of 8898 records. For pain intensity, a mean difference of (MD: -5.32, 95% confidence interval (CI): -7.20, -3.44, N = 57 studies with 74 outcomes, GRADE: very low) was estimated for placebo interventions. A small effect in favour of the placebo interventions for physical function was estimated (SMD: -0.22, 95% CI: -0.35, -0.09; N = 37 with 48 outcomes, GRADE: very low). Similar results were found for a broad range of patient-reported outcomes. Meta-regression analyses did not explain heterogeneity among analyses. CONCLUSION: The study found that the contextual effect of non-pharmacological conservative interventions for musculoskeletal conditions is likely to be small. However, given the known effect sizes of recommended evidence-based treatments for musculoskeletal conditions, it may still contribute an important component. SIGNIFICANCE: Contextual effects of non-pharmacological conservative interventions for musculoskeletal conditions are likely to be small for a broad range of patient-reported outcomes (pain intensity, physical function, quality of life, global rating of change and depression). Contextual effects are unlikely, in isolation, to offer much clinical care. But these factors do have relevance in an overall treatment context as they provide almost 30% of the minimally clinically important difference.

Integrative single-cell meta-analysis reveals disease-relevant vascular cell states and markers in human atherosclerosis.

Coronary artery disease (CAD) is characterized by atherosclerotic plaque formation in the arterial wall. CAD progression involves complex interactions and phenotypic plasticity among vascular and immune cell lineages. Single-cell RNA-seq (scRNA-seq) studies have highlighted lineage-specific transcriptomic signatures, but human cell phenotypes remain controversial. Here, we perform an integrated meta-analysis of 22 scRNA-seq libraries to generate a comprehensive map of human atherosclerosis with 118,578 cells. Besides characterizing granular cell-type diversity and communication, we leverage this atlas to provide insights into smooth muscle cell (SMC) modulation. We integrate genome-wide association study data and uncover a critical role for modulated SMC phenotypes in CAD, myocardial infarction, and coronary calcification. Finally, we identify fibromyocyte/fibrochondrogenic SMC markers (LTBP1 and CRTAC1) as proxies of atherosclerosis progression and validate these through omics and spatial imaging analyses. Altogether, we create a unified atlas of human atherosclerosis informing cell state-specific mechanistic and translational studies of cardiovascular diseases.

Gene-educational attainment interactions in a multi-population genome-wide meta-analysis identify novel lipid loci.

Introduction: Educational attainment, widely used in epidemiologic studies as a surrogate for socioeconomic status, is a predictor of cardiovascular health outcomes. Methods: A two-stage genome-wide meta-analysis of low-density lipoprotein cholesterol (LDL), high-density lipoprotein cholesterol (HDL), and triglyceride (TG) levels was performed while accounting for gene-educational attainment interactions in up to 226,315 individuals from five population groups. We considered two educational attainment variables: "Some College" (yes/no, for any education beyond high school) and "Graduated College" (yes/no, for completing a 4-year college degree). Genome-wide significant (p < 5 × 10-8) and suggestive (p < 1 × 10-6) variants were identified in Stage 1 (in up to 108,784 individuals) through genome-wide analysis, and those variants were followed up in Stage 2 studies (in up to 117,531 individuals). Results: In combined analysis of Stages 1 and 2, we identified 18 novel lipid loci (nine for LDL, seven for HDL, and two for TG) by two degree-of-freedom (2 DF) joint tests of main and interaction effects. Four loci showed significant interaction with educational attainment. Two loci were significant only in cross-population analyses. Several loci include genes with known or suggested roles in adipose (FOXP1, MBOAT4, SKP2, STIM1, STX4), brain (BRI3, FILIP1, FOXP1, LINC00290, LMTK2, MBOAT4, MYO6, SENP6, SRGAP3, STIM1, TMEM167A, TMEM30A), and liver (BRI3, FOXP1) biology, highlighting the potential importance of brain-adipose-liver communication in the regulation of lipid metabolism. An investigation of the potential druggability of genes in identified loci resulted in five gene targets shown to interact with drugs approved by the Food and Drug Administration, including genes with roles in adipose and brain tissue. Discussion: Genome-wide interaction analysis of educational attainment identified novel lipid loci not previously detected by analyses limited to main genetic effects.

Treatment of calcific arterial disease via enhancement of autophagy using GSK343.

Vascular calcification is a hallmark of atherosclerotic disease and serves as a strong predictor and risk factor for cardiovascular events. Growing evidence suggests that autophagy may play a protective role in early atherosclerosis. The precise effects of autophagy on VSMC-mediated calcification remain unknown. In this study, we utilized multi-omic profiling to investigate impaired autophagy at the transcriptional level as a key driver of VSMC calcification. Our findings revealed that impaired autophagy is an essential determinant of VSMC calcification. We observed that an osteogenic environment affects the open chromatin status of VSMCs, compromising the transcriptional activation of autophagy initiation genes. In vivo experiments involve pharmacological and genetic activation of autophagy using mouse models of spontaneous large (Mgp-/-) and small (Abcc6-/-) artery calcification. Taken together, these data advance our mechanistic understanding of vascular calcification and provide important insights for a broad range of cardiovascular diseases involving VSMC phenotype switch.

The accumulation of erythrocytes quantified and visualized by Glycophorin C in carotid atherosclerotic plaque reflects intraplaque hemorrhage and pre-procedural neurological symptoms.

The accumulation of erythrocyte membranes within an atherosclerotic plaque may contribute to the deposition of free cholesterol and thereby the enlargement of the necrotic core. Erythrocyte membranes can be visualized and quantified in the plaque by immunostaining for the erythrocyte marker glycophorin C. Hence, we theorized that the accumulation of erythrocytes quantified by glycophorin C could function as a marker for plaque vulnerability, possibly reflecting intraplaque hemorrhage (IPH), and offering predictive value for pre-procedural neurological symptoms. We employed the CellProfiler-integrated slideToolKit workflow to visualize and quantify glycophorin C, defined as the total plaque area that is positive for glycophorin C, in single slides of culprit lesions obtained from the Athero-Express Biobank of 1819 consecutive asymptomatic and symptomatic patients who underwent carotid endarterectomy. Our assessment included the evaluation of various parameters such as lipid core, calcifications, collagen content, SMC content, and macrophage burden. These parameters were evaluated using a semi-quantitative scoring method, and the resulting data was dichotomized as predefined criteria into categories of no/minor or moderate/heavy staining. In addition, the presence or absence of IPH was also scored. The prevalence of IPH and pre-procedural neurological symptoms were 62.4% and 87.1%, respectively. The amount of glycophorin staining was significantly higher in samples from men compared to samples of women (median 7.15 (IQR:3.37, 13.41) versus median 4.06 (IQR:1.98, 8.32), p < 0.001). Glycophorin C was associated with IPH adjusted for clinical confounders (OR 1.90; 95% CI 1.63, 2.21; p = < 0.001). Glycophorin C was significantly associated with ipsilateral pre-procedural neurological symptoms (OR:1.27, 95%CI:1.06-1.41, p = 0.005). Sex-stratified analysis, showed that this was also the case for men (OR 1.37; 95%CI 1.12, 1.69; p = 0.003), but not for women (OR 1.15; 95%CI 0.77, 1.73; p = 0.27). Glycophorin C was associated with classical features of a vulnerable plaque, such as a larger lipid core, a higher macrophage burden, less calcifications, a lower collagen and SMC content. There were marked sex differences, in men, glycophorin C was associated with calcifications and collagen while these associations were not found in women. To conclude, the accumulation of erythrocytes in atherosclerotic plaque quantified and visualized by glycophorin C was independently associated with the presence of IPH, preprocedural symptoms in men, and with a more vulnerable plaque composition in both men and women. These results strengthen the notion that the accumulation of erythrocytes quantified by glycophorin C can be used as a marker for plaque vulnerability.

Evidence- and data-driven classification of low back pain via artificial intelligence: Protocol of the PREDICT-LBP study.

In patients presenting with low back pain (LBP), once specific causes are excluded (fracture, infection, inflammatory arthritis, cancer, cauda equina and radiculopathy) many clinicians pose a diagnosis of non-specific LBP. Accordingly, current management of non-specific LBP is generic. There is a need for a classification of non-specific LBP that is both data- and evidence-based assessing multi-dimensional pain-related factors in a large sample size. The "PRedictive Evidence Driven Intelligent Classification Tool for Low Back Pain" (PREDICT-LBP) project is a prospective cross-sectional study which will compare 300 women and men with non-specific LBP (aged 18-55 years) with 100 matched referents without a history of LBP. Participants will be recruited from the general public and local medical facilities. Data will be collected on spinal tissue (intervertebral disc composition and morphology, vertebral fat fraction and paraspinal muscle size and composition via magnetic resonance imaging [MRI]), central nervous system adaptation (pain thresholds, temporal summation of pain, brain resting state functional connectivity, structural connectivity and regional volumes via MRI), psychosocial factors (e.g. depression, anxiety) and other musculoskeletal pain symptoms. Dimensionality reduction, cluster validation and fuzzy c-means clustering methods, classification models, and relevant sensitivity analyses, will classify non-specific LBP patients into sub-groups. This project represents a first personalised diagnostic approach to non-specific LBP, with potential for widespread uptake in clinical practice. This project will provide evidence to support clinical trials assessing specific treatments approaches for potential subgroups of patients with non-specific LBP. The classification tool may lead to better patient outcomes and reduction in economic costs.

Towards data-driven biopsychosocial classification of non-specific chronic low back pain: a pilot study.

The classification of non-specific chronic low back pain (CLBP) according to multidimensional data could guide clinical management; yet recent systematic reviews show this has not been attempted. This was a prospective cross-sectional study of participants with CLBP (n = 21) and age-, sex- and height-matched pain-free controls (n = 21). Nervous system, lumbar spinal tissue and psychosocial factors were collected. Dimensionality reduction was followed by fuzzy c-means clustering to determine sub-groups. Machine learning models (Support Vector Machine, k-Nearest Neighbour, Naïve Bayes and Random Forest) were used to determine the accuracy of classification to sub-groups. The primary analysis showed that four factors (cognitive function, depressive symptoms, general self-efficacy and anxiety symptoms) and two clusters (normal versus impaired psychosocial profiles) optimally classified participants. The error rates in classification models ranged from 4.2 to 14.2% when only CLBP patients were considered and increased to 24.2 to 37.5% when pain-free controls were added. This data-driven pilot study classified participants with CLBP into sub-groups, primarily based on psychosocial factors. This contributes to the literature as it was the first study to evaluate data-driven machine learning CLBP classification based on nervous system, lumbar spinal tissue and psychosocial factors. Future studies with larger sample sizes should validate these findings.