Major Facilitator Superfamily Domain Containing 5 Inhibition Reduces Lipoprotein(a) Uptake and Calcification in Valvular Heart Disease.

BACKGROUND: High circulating levels of Lp(a) (lipoprotein[a]) increase the risk of atherosclerosis and calcific aortic valve disease, affecting millions of patients worldwide. Although atherosclerosis is commonly treated with low-density lipoprotein-targeting therapies, these do not reduce Lp(a) or risk of calcific aortic valve disease, which has no available drug therapies. Targeting Lp(a) production and catabolism may provide therapeutic benefit, but little is known about Lp(a) cellular uptake. METHODS: Here, unbiased ligand-receptor capture mass spectrometry was used to identify MFSD5 (major facilitator superfamily domain containing 5) as a novel receptor/cofactor involved in Lp(a) uptake. RESULTS: Reducing MFSD5 expression by a computationally identified small molecule or small interfering RNA suppressed Lp(a) uptake and calcification in primary human valvular endothelial and interstitial cells. MFSD5 variants were associated with aortic stenosis (P=0.027 after multiple hypothesis testing) with evidence suggestive of an interaction with plasma Lp(a) levels. CONCLUSIONS: MFSD5 knockdown suppressing human valvular cell Lp(a) uptake and calcification, along with meta-analysis of MFSD5 variants associating with aortic stenosis, supports further preclinical assessment of MFSD5 in cardiovascular diseases, the leading cause of death worldwide.

Lipoprotein(a) and carotid intima-media thickness in children with familial hypercholesterolaemia in the Netherlands: a 20-year follow-up study.

BACKGROUND: Elevated lipoprotein(a) and familial hypercholesterolaemia are both independent risk conditions for cardiovascular disease. Although signs of atherosclerosis can be observed in children with familial hypercholesterolaemia, it is unknown whether elevated lipoprotein(a) is an additional risk factor for atherosclerosis in these young patients. Therefore, we aimed to assess the contribution of lipoprotein(a) concentrations to arterial wall thickening (as measured by carotid intima-media thickness) in children with familial hypercholesterolaemia who were followed up into adulthood. METHODS: We conducted a 20-year follow-up study of 214 children (aged 8-18 years) with heterozygous familial hypercholesterolaemia who were randomly assigned in a statin trial in Amsterdam (Netherlands) between Dec 7, 1997, and Oct 4, 1999. At baseline, and at 2, 10, and 20 years thereafter, blood samples were taken and carotid intima-media thickness was measured. Linear mixed-effects models were used to evaluate the association between lipoprotein(a) and carotid intima-media thickness during follow-up. We adjusted for sex, age, corrected LDL-cholesterol, statin use, and BMI. FINDINGS: Our study population comprised 200 children who had a carotid intima-media thickness measurement and a measured lipoprotein(a) concentration from at least one visit available. Mean age at baseline was 13·0 years (SD 2·9), 106 (53%) children were male, and 94 (47%) were female. At baseline, median lipoprotein(a) concentration was 18·5 nmol/L (IQR 8·7-35·5) and mean carotid intima-media thickness was 0·4465 mm (SD 0·0496). During follow-up, higher lipoprotein(a) concentrations contributed significantly to progression of carotid intima-media thickness (ß adjusted 0·0073 mm per 50 nmol/L increase in lipoprotein(a) [95% CI 0·0013-0·0132]; p=0·017). INTERPRETATION: Our findings suggest that lipoprotein(a) concentrations contribute significantly to arterial wall thickening in children with familial hypercholesterolaemia who were followed-up until adulthood, suggesting that lipoprotein(a) is an independent and additional risk factor for early atherosclerosis in those already at increased risk. Lipoprotein(a) measurement in young patients with familial hypercholesterolaemia is crucial to identify those at potentially highest risk for cardiovascular disease. FUNDING: Silence Therapeutics.

DNA methyltransferase 3 alpha and TET methylcytosine dioxygenase 2 restrain mitochondrial DNA-mediated interferon signaling in macrophages.

Deleterious somatic mutations in DNA methyltransferase 3 alpha (DNMT3A) and TET mehtylcytosine dioxygenase 2 (TET2) are associated with clonal expansion of hematopoietic cells and higher risk of cardiovascular disease (CVD). Here, we investigated roles of DNMT3A and TET2 in normal human monocyte-derived macrophages (MDM), in MDM isolated from individuals with DNMT3A or TET2 mutations, and in macrophages isolated from human atherosclerotic plaques. We found that loss of function of DNMT3A or TET2 resulted in a type I interferon response due to impaired mitochondrial DNA integrity and activation of cGAS signaling. DNMT3A and TET2 normally maintained mitochondrial DNA integrity by regulating the expression of transcription factor A mitochondria (TFAM) dependent on their interactions with RBPJ and ZNF143 at regulatory regions of the TFAM gene. These findings suggest that targeting the cGAS-type I IFN pathway may have therapeutic value in reducing risk of CVD in patients with DNMT3A or TET2 mutations.

Clonal hematopoiesis driven by DNMT3A and TET2 mutations: role in monocyte and macrophage biology and atherosclerotic cardiovascular disease.

PURPOSE OF REVIEW: Clonal hematopoiesis of indeterminate potential (CHIP), defined by the presence of somatic mutations in hematopoietic cells, is associated with advanced age and increased mortality due to cardiovascular disease. Gene mutations in DNMT3A and TET2 are the most frequently identified variants among patients with CHIP and provide selective advantage that spurs clonal expansion and myeloid skewing. Although DNMT3A and TET2 appear to have opposing enzymatic influence on DNA methylation, mounting data has characterized convergent inflammatory pathways, providing insights to how CHIP may mediate atherosclerotic cardiovascular disease (ASCVD). RECENT FINDINGS: We review a multitude of studies that characterize aberrant inflammatory signaling as result of DNMT3A and TET2 deficiency in monocytes and macrophages, immune cells with prominent roles in atherosclerosis. Although specific DNA methylation signatures associated with these known epigenetic regulators have been identified, many studies have also characterized diverse modulatory functions of DNTM3A and TET2 that urge cell and context-specific experimental studies to further define how DNMT3A and TET2 may nonenzymatically activate inflammatory pathways with clinically meaningful consequences. SUMMARY: CHIP, common in elderly individuals, provides an opportunity understand and potentially modify age-related chronic inflammatory ASCVD risk.

The interconnection between lipoprotein(a), lipoprotein(a) cholesterol and true LDL-cholesterol in the diagnosis of familial hypercholesterolemia.

PURPOSE OF REVIEW: Elevated levels of lipoprotein(a) [Lp(a)] are present in 30-50% of patients with familial hypercholesterolemia. The contribution of Lp(a) towards risk stratification of patients with familial hypercholesterolemia has been recently recognized, with studies showing a significantly worse prognosis if Lp(a) is elevated. However, the role of elevated Lp(a) in diagnosis of familial hypercholesterolemia is less well defined or accepted. RECENT FINDINGS: An important confounder in the diagnosis of familial hypercholesterolemia is the significant contribution of the cholesterol content on Lp(a) (Lp(a)-C) in individuals with elevated Lp(a). Because Lp(a)-C is incorporated into all clinical LDL-C measurements, it can contribute significantly to the cholesterol threshold diagnostic criteria for familial hypercholesterolemia used in most clinical algorithms. SUMMARY: In this review, we discuss the interrelationship of Lp(a), Lp(a)-C and correct LDL-C in the diagnosis and prognosis of familial hypercholesterolemia. Future studies of accurately measuring correct LDL-C or in using apoB-100 and Lp(a) criteria may overcome the limitations of using estimated LDL-C in the diagnosis of familial hypercholesterolemia in individuals with concomitant elevation of Lp(a).

Short-term regulation of hematopoiesis by lipoprotein(a) results in the production of pro-inflammatory monocytes.

BACKGROUND: Lipoproteins are important regulators of hematopoietic stem and progenitor cell (HSPC) biology, predominantly affecting myelopoiesis. Since myeloid cells, including monocytes and macrophages, promote the inflammatory response that propagates atherosclerosis, it is of interest whether the atherogenic low-density lipoprotein (LDL)-like particle lipoprotein(a) [Lp(a)] contributes to atherogenesis via stimulating myelopoiesis. METHODS & RESULTS: To assess the effects of Lp(a)-priming on long-term HSPC behavior we transplanted BM of Lp(a) transgenic mice, that had been exposed to elevated levels of Lp(a), into lethally-irradiated C57Bl6 mice and hematopoietic reconstitution was analyzed. No differences in HSPC populations or circulating myeloid cells were detected ten weeks after transplantation. Likewise, in vitro stimulation of C57Bl6 BM cells for 24 h with Lp(a) did not affect colony formation, total cell numbers or myeloid populations 7 days later. To assess the effects of elevated levels of Lp(a) on myelopoiesis, C57Bl6 bone marrow (BM) cells were stimulated with lp(a) for 24 h, and a marked increase in granulocyte-monocyte progenitors, pro-inflammatory Ly6high monocytes and macrophages was observed. Seven days of continuous exposure to Lp(a) increased colony formation and enhanced the formation of pro-inflammatory monocytes and macrophages. Antibody-mediated neutralization of oxidized phospholipids abolished the Lp(a)-induced effects on myelopoiesis. CONCLUSION: Lp(a) enhances the production of inflammatory monocytes at the bone marrow level but does not induce cell-intrinsic long-term priming of HSPCs. Given the short-term and direct nature of this effect, we postulate that Lp(a)-lowering treatment has the capacity to rapidly revert this multi-level inflammatory response.

ApoCIII-Lp(a) complexes in conjunction with Lp(a)-OxPL predict rapid progression of aortic stenosis.

OBJECTIVE: This study assessed whether apolipoprotein CIII-lipoprotein(a) complexes (ApoCIII-Lp(a)) associate with progression of calcific aortic valve stenosis (AS). METHODS: Immunostaining for ApoC-III was performed in explanted aortic valve leaflets in 68 patients with leaflet pathological grades of 1-4. Assays measuring circulating levels of ApoCIII-Lp(a) complexes were measured in 218 patients with mild-moderate AS from the AS Progression Observation: Measuring Effects of Rosuvastatin (ASTRONOMER) trial. The progression rate of AS, measured as annualised changes in peak aortic jet velocity (Vpeak), and combined rates of aortic valve replacement (AVR) and cardiac death were determined. For further confirmation of the assay data, a proteomic analysis of purified Lp(a) was performed to confirm the presence of apoC-III on Lp(a). RESULTS: Immunohistochemically detected ApoC-III was prominent in all grades of leaflet lesion severity. Significant interactions were present between ApoCIII-Lp(a) and Lp(a), oxidised phospholipids on apolipoprotein B-100 (OxPL-apoB) or on apolipoprotein (a) (OxPL-apo(a)) with annualised Vpeak (all p<0.05). After multivariable adjustment, patients in the top tertile of both apoCIII-Lp(a) and Lp(a) had significantly higher annualised Vpeak (p<0.001) and risk of AVR/cardiac death (p=0.03). Similar results were noted with OxPL-apoB and OxPL-apo(a). There was no association between autotaxin (ATX) on ApoB and ATX on Lp(a) with faster progression of AS. Proteomic analysis of purified Lp(a) showed that apoC-III was prominently present on Lp(a). CONCLUSION: ApoC-III is present on Lp(a) and in aortic valve leaflets. Elevated levels of ApoCIII-Lp(a) complexes in conjunction with Lp(a), OxPL-apoB or OxPL-apo(a) identify patients with pre-existing mild-moderate AS who display rapid progression of AS and higher rates of AVR/cardiac death. TRIAL REGISTRATION: NCT00800800.

Lipoprotein(a) in Patients Undergoing Transcatheter Aortic Valve Replacement.

Lipoprotein(a) [Lp(a)] is a genetically determined risk factor for calcific aortic valve stenosis (CAVS) for which transcatheter aortic valve replacement (TAVR) is increasingly utilized as treatment. We evaluated the effect of a program to increase testing of and define the prevalence of elevated Lp(a) among patients undergoing TAVR. Educational efforts and incorporation of a "check-box" Lp(a) order to the preoperative TAVR order set were instituted. Retrospective chart review was performed in 229 patients requiring TAVR between May 2013 and September 2018. Of these patients, 57% had an Lp(a) level measured; testing rates increased from 0% in 2013 to 96% in 2018. Lipoprotein(a) testing occurred in 11% of patients before and in 80% of patients after the "check-box" order set ( P < .001). The prevalence of elevated Lp(a) (≥30 mg/dL) was 35%; these patients had a higher incidence of coronary artery disease requiring revascularization compared with patients with normal Lp(a) (65% vs 47%; P = .047). Patients with Lp(a) ≥30 mg/dL also had higher incidence of paravalvular leak compared with those with normal Lp(a) (13% vs 4%; P = .04). This study defines the prevalence of elevated Lp(a) in advanced stages of CAVS and provides a practice pathway to assess procedural complications and long-term outcomes of TAVR in patients with elevated Lp(a) levels.

The Prevalence of Lipoprotein(a) Measurement and Degree of Elevation Among 2710 Patients With Calcific Aortic Valve Stenosis in an Academic Echocardiography Laboratory Setting.

Lipoprotein(a; Lp[a]) and its associated oxidized phospholipids are causal, genetic risk factors for calcific aortic valve stenosis (CAVS). We determined the prevalence of Lp(a) measurement among 2710 patients with CAVS and 1369 control patients (∼50% of study group) without CAVS with an echocardiogram between January 2010 and February 2016 in an academic echocardiography laboratory. Lipoprotein(a) measurements were performed at a referral laboratory using an isoform-independent assay. The prevalence of any Lp(a) measurement was 4.6% (124 of the 2710) in patients with CAVS and 3.1% (42 of the 1369) in the control group ( P = .021). In patients with CAVS, mean (standard deviation) Lp(a) levels were 38 (54) mg/dL and median (interquartile range) Lp(a) levels were 14 (6-48) mg/dL. Of the 124 patients with CAVS having Lp(a) measurements, 83 (66.9%) had Lp(a) <30 mg/dL and 41 (33.1%) had Lp(a) ≥30 mg/dL. This study reflects low physician testing of Lp(a) levels in CAVS. Given the role of Lp(a) as a causal risk factor for CAVS, and the ongoing development of therapies to normalize Lp(a) levels, our results suggest that Lp(a) measurements in CAVS should be more widely obtained in clinical practice.

Lipoprotein(a) and oxidized phospholipids in calcific aortic valve stenosis.

PURPOSE OF REVIEW: As the incidence of calcific aortic valve stenosis increases with the aging of the population, improved understanding and novel therapies to reduce its progression and need for aortic valve replacement are urgently needed. RECENT FINDINGS: Lipoprotein(a) is the only monogenetic risk factor for calcific aortic stenosis. Elevated levels are a strong, causal, independent risk factor, as demonstrated in epidemiological, genome-wide association studies and Mendelian randomization studies. Lipoprotein(a) is the major lipoprotein carrier of oxidized phospholipids, which are proinflammatory and promote calcification of vascular cells, two key pathophysiological drivers of aortic stenosis. Elevated plasma lipoprotein(a) and oxidized phospholipids predict progression of pre-existing aortic stenosis and need for aortic valve replacement. The failure of statin trials in pre-existing aortic stenosis may be partially due to an increase in lipoprotein(a) and oxidized phospholipid levels caused by statins. Antisense oligonucleotides targeted to apo(a) are in Phase 2 clinical development and shown to lower both lipoprotein(a) and oxidized phospholipids. SUMMARY: Lipoprotein(a) and oxidized phospholipids are key therapeutic targets in calcific aortic stenosis. Strategies aimed at potent lipoprotein(a) lowering to normalize levels and/or to suppress the proinflammatory effects of oxidized phospholipids may prevent progression of this disease.

Oxidized Phospholipids, Lipoprotein(a), and Progression of Calcific Aortic Valve Stenosis.

BACKGROUND: Elevated lipoprotein(a) (Lp[a]) is associated with aortic stenosis (AS). Oxidized phospholipids (OxPL) are key mediators of calcification in valvular cells and are carried by Lp(a). OBJECTIVES: This study sought to determine whether Lp(a) and OxPL are associated with hemodynamic progression of AS and AS-related events. METHODS: OxPL on apolipoprotein B-100 (OxPL-apoB), which reflects the biological activity of Lp(a), and Lp(a) levels were measured in 220 patients with mild-to-moderate AS. The primary endpoint was the progression rate of AS, measured by the annualized increase in peak aortic jet velocity in m/s/year by Doppler echocardiography; the secondary endpoint was need for aortic valve replacement and cardiac death during 3.5 ± 1.2 years of follow-up. RESULTS: AS progression was faster in patients in the top tertiles of Lp(a) (peak aortic jet velocity: +0.26 ± 0.26 vs. +0.17 ± 0.21 m/s/year; p = 0.005) and OxPL-apoB (+0.26 ± 0.26 m/s/year vs. +0.17 ± 0.21 m/s/year; p = 0.01). After multivariable adjustment, elevated Lp(a) or OxPL-apoB levels remained independent predictors of faster AS progression. After adjustment for age, sex, and baseline AS severity, patients in the top tertile of Lp(a) or OxPL-apoB had increased risk of aortic valve replacement and cardiac death. CONCLUSIONS: Elevated Lp(a) and OxPL-apoB levels are associated with faster AS progression and need for aortic valve replacement. These findings support the hypothesis that Lp(a) mediates AS progression through its associated OxPL and provide a rationale for randomized trials of Lp(a)-lowering and OxPL-apoB-lowering therapies in AS. (Aortic Stenosis Progression Observation: Measuring Effects of Rosuvastatin [ASTRONOMER]; NCT00800800).

Role of phospholipid transfer protein in high-density lipoprotein- mediated reverse cholesterol transport.

Reverse cholesterol transport (RCT) describes the process whereby cholesterol in peripheral tissues is transported to the liver where it is ultimately excreted in the form of bile. Given the atherogenic role of cholesterol accumulation within the vessel intima, removal of cholesterol through RCT is considered an anti-atherogenic process. The major constituents of RCT include cell membrane- bound lipid transporters, plasma lipid acceptors, plasma proteins and enzymes, and lipid receptors of liver cell membrane. One major cholesterol acceptor in RCT is high-density lipoprotein (HDL). Both the characteristics and level of HDL are critical determinants for RCT. It is known that phospholipid transfer protein (PLTP) impacts both HDL cholesterol level and biological quality of the HDL molecule. Recent data suggest that PLTP has a site-specific variation in its function. Moreover, the RCT pathway also has multiple steps both in the peripheral tissues and circulation. Therefore, PLTP may influence the RCT pathway at multiple levels. In this review, we focus on the potential role of PLTP in RCT through its impact on HDL homeostasis. The relationship between PLTP and RCT is expected to be an important area in finding novel therapies for atherosclerosis.

Sphingomyelin synthase 2 (SMS2) deficiency attenuates LPS-induced lung injury.

Sphingomyelin synthase (SMS) catalyzes the synthesis of sphingomyelin (SM) and is required for maintenance of plasma membrane microdomain fluidity. Of the two isoforms of mammalian SMS, SMS1 is mostly present in the trans-Golgi apparatus, whereas SMS2 is predominantly found at the plasma membrane. SMS2 has a role in receptor mediated response to inflammation in macrophages, however, the role of SMS2 in vascular permeability, pulmonary edema, and lung injury have not been investigated. To define the role of SMS activation in lung injury, we utilized a lipopolysaccharide (LPS)-induced lung edema model. SMS activity was measured and correlated with the severity of lung injury. Within 4 h of LPS treatment, SMS activity was increased significantly and remained upregulated up to 24 h. Comparison of LPS-induced lung injury in SMS2 knockout (SMS2(-/-)) and wild-type littermate control mice showed that inflammation, cytokine induction, and lung injury were significantly inhibited in SMS2(-/-) mice. Our results suggest that a deficiency of SMS2 can diminish the extent of pulmonary edema and lung injury. Furthermore, we show that depletion of SMS2 was sufficient to decrease MAP kinase-JNK activation, severity of LPS-induced pulmonary neutrophil influx, and inflammation, suggesting a novel role of SMS2 activation in lung injury.