Ileana Badi

BACKGROUND: Lp(a) (lipoprotein[a]) is associated with cardiovascular disease, but neither the causal nature nor the underlying mechanisms are fully documented. This study investigated whether Lp(a) triggers atherogenesis by dysregulating vascular redox–sensitive inflammatory state. METHODS: Plasma Lp(a) was measured in 1027 patients with advanced coronary artery disease undergoing cardiac surgery. These patients were genotyped, and a modified LPA genetic risk score (LPA GRS) determining Lp(a) levels was generated. RNA sequencing and vascular superoxide measurements were performed in internal mammary arteries, and the contribution of NOXs (NADPH oxidases) and uncoupled eNOS (endothelial nitric oxide synthase) was determined. The median follow-up was 5.07 years. RESULTS: Increased plasma Lp(a) ( P =0.03) and LPA GRS ( P =0.01) were associated with elevated arterial superoxide in the overall patient population, an effect that was driven by nondiabetics. This effect was primarily due to eNOS uncoupling via reduced vascular BH4 (tetrahydrobiopterin) bioavailability. There was no significant impact of Lp(a) variability on vascular NOX–derived superoxide ( P =0.13). RNA sequencing of arterial tissue revealed dysregulation of nitrosative and inflammatory signaling in high Lp(a) patients although there was no association with systemic biomarkers of inflammation (ie, hsCRP [high-sensitivity C-reactive protein]; P =0.82) or oxidative stress (ie, malondialdehyde; P =0.61). Finally, both LPA GRS (hazard ratio, 3.615 [95% CI, 1.044–12.515]; P =0.043) and high plasma Lp(a) (hazard ratio, 3.286 [95% CI, 1.003–10.767]; P =0.049) were associated with elevated risk for cardiac mortality. This association was vascular superoxide-dependent, implying that redox-sensitive inflammatory signaling may be a link between Lp(a) and cardiovascular risk. All the above associations were independent of plasma ApoB (apolipoprotein-B). CONCLUSIONS: This study demonstrates for the first time that a genetically determined increase in plasma Lp(a) results in dysregulated vascular redox/nitrosative signaling in patients with atherosclerosis.

Background: Visceral obesity is directly linked to increased cardiovascular risk, including heart failure. Purpose: We explored the ability of human epicardial adipose tissue (EAT)-derived microRNAs (miRNAs) to regulate the myocardial redox state and clinical outcomes. Methods: The 5 study arms included 466 patients undergoing cardiac surgery to perform: 1) the discovery phase in which we screened for 351 miRNAs expressed and released from human EAT; 2) correlation analyses between EAT microRNAs and myocardial expression of their targets or myocardial superoxide production in paired EAT/atrial biopsies; 3) genome-wide association screening for miR-92a-3p expression in EAT to test causality/directionality; 4) ex vivo experiments to investigate the underlying mechanisms that were also studied in vitro and in vivo; 5) an 8-year follow-up study to test the prognostic value of the discovered miRNA. Superoxide (O2.-) generation was measured by lucigenin chemiluminescence with NADPH 100μM stimulation as indicator of NADPH-oxidases activity. Vas2870 400 μM (a specific pan-NADPH oxidase inhibitor) was used to obtain the Vas2870-inhibitable O2.- signal which constitutes a more specific index of NADPH oxidase activity. Activation of Rac1, a key NADPH-oxidases subunit, was evaluated by a commercially available kit. Differentiated H9c2 cells were used as an in vitro model of cardiomyocytes. Doxycycline-inducible Wnt5a-overexpressing mice were used for in vivo experiments. Results: The EAT secretome profiling on study 1 patients led us to identify 3 microRNAs both expressed and released by EAT whose levels in EAT correlated with oxidative stress in human myocardium. Among these miRNAs only miR-92a-3p reduced NADPH-oxidase-derived superoxide (O2.-) in cardiomyocytes (A). A genetic screening identified 7 SNPs that were associated with high miR-92a-3p levels in EAT (EAT-miR-92a-3p SNPs) and were related to lower myocardial superoxide production (B). miR-92a-3p decreased both activation of Rac1 (not shown) and Wnt5a protein levels in vitro (C). Patients with high EAT miR-92a-3p levels had lower WNT5A levels in the myocardium (not shown). Ex vivo, in vivo and in vitro experiments showed opposite effects of Wnt5a on Rac1-mediated NADPH-oxidases activity (D and not shown). Finally, we found an association of miR-92a-3p levels in EAT with lower relative risk of adverse cardiovascular events (E). Conclusions: EAT-derived miRNAs exert paracrine effects on the human heart. Indeed miR-92a-3p suppresses the Wnt5a/Rac1/NADPH oxidase axis and improves myocardial redox state. EAT-derived miR-92a-3p is related with improved clinical outcomes and is a rational therapeutic target for the prevention and treatment of obesity-related heart disease.

Introduction Lp(a) levels has been associated with increased cardiovascular risk and is a therapeutic target in cardiovascular prevention. However, the mechanisms behind this association are unclear. A new radiotranscriptomic library (the oxHVF platform) uses RNA sequencing data from human arteries and perivascular adipose tissue (PVAT) as ground truth for the development of machine-learning powered imaging biomarkers of vascular biology. We aimed to use this radiotranscriptomic platform to understand the mechanisms by which Lp(a) affects vascular function in patients with advanced atherosclerosis. Methods A cohort of 1472 patients undergoing CABG surgery were recruited. Plasma Lp(a) levels were measured before surgery. We performed genotyping to identify a combination of the top 7 SNPs that are associated with high Lp(a) levels. Internal mammary arteries (IMA) segments were obtained to quantify ex vivo superoxide generation and phenotype vascular inflammation. 369 of these patients underwent CCTA, and PVAT inflammation in the IMA and the aorta was measured using a recently developed radiotranscriptomic signature of vascular cytokine-driven inflammation (C19-RS) (A). A subgroup of 50 patients had CCTAs at 6 months and 5 years after surgery, to access progression of SVG disease. Results Patients with high plasma Lp(a) levels (>95th centile, >106mg/dL) had higher vascular inflammation measured by C19RS (B and C). Similarly, patients carrying a genetic signature associated with high Lp(a) levels also experienced high C19RS (D), suggesting a causal association. Patients with high Lp(a) (E) or genetically determined Lp(a) elevation (F) also had higher O2- generation in their IMAs (E, F). In the subgroup with paired CCTA scans, high Lp(a) levels at baseline had significantly greater changes in SVG intimal hyperplasia compared with those with low Lp(a) levels (G). Conclusions High Lp(a) levels and a genetic signature driving plasma Lp(a), are related with vascular redox dysregulation, resulting into increased vascular inflammation measured by using CCTA radiotranscriptomic phenotyping (C19RS), in patients with advanced atherosclerosis. Moreover, Lp(a) levels predict the development of intima hyperplasia in the SVGs of the same patients, implying involvement in atherosclerotic disease progression. Conflict of Interest Nil

Editorial: Exploring the Crosstalk Between Adipose Tissue and the Cardiovascular System

The senescence of vascular smooth muscle cells (VSMCs), characterized by the acquisition of senescence-associated secretory phenotype (SASP), is relevant for VSMCs osteoblastic differentiation and vascular calcification (VC). MicroRNA-34a (miR-34a) is a driver of such phenomena and could play a role in vascular inflammaging. Herein, we analyzed the relationship between miR-34a and the prototypical SASP component IL6 in in vitro and in vivo models. miR-34a and IL6 levels increased and positively correlated in aortas of 21 months-old male C57BL/6J mice and in human aortic smooth muscle cells (HASMCs) isolated from donors of different age and undergone senescence. Lentiviral overexpression of miR-34a in HASMCs enhanced IL6 secretion. HASMCs senescence and calcification accelerated after exposure to conditioned medium of miR-34a-overexpressing cells. Analysis of miR-34a-induced secretome revealed enhancement of several pro-inflammatory cytokines and chemokines, including IL6, pro-senescent growth factors and matrix-degrading molecules. Moreover, induction of aortas medial calcification and concomitant IL6 expression, with an overdose of vitamin D, was reduced in male C57BL/6J Mir34a^−/− mice. Finally, a positive correlation was observed between circulating miR-34a and IL6 in healthy subjects of 20-90 years. Hence, the vascular age-associated miR-34a promotes VSMCs SASP activation and contributes to arterial inflammation and dysfunctions such as VC.