SPAG5 deficiency activates autophagy to reduce atherosclerotic plaque formation in ApoE-/- mice.

BACKGROUND: Autophagy, as a regulator of cell survival, plays an important role in atherosclerosis (AS). Sperm associated antigen 5 (SPAG5) is closely associated with the classical autophagy pathway, PI3K/Akt/mTOR signaling pathway. This work attempted to investigate whether SPAG5 can affect AS development by regulating autophagy. METHODS: Human umbilical vein endothelial cells (HUVECs) were treated with oxidized-low density lipoprotein (ox-LDL) to induce cell damage. ApoE-/- mice were fed a Western diet to establish an AS mouse model. Haematoxylin and eosin (H&E) staining and Oil Red O staining evaluated the pathological changes and in lipid deposition in aortic tissues. CCK-8 and flow cytometry detected cell proliferation and apoptosis. Immunohistochemistry, Enzyme linked immunosorbent assay, qRT-PCR and western blotting assessed the levels of mRNA and proteins. RESULTS: Ox-LDL treatment elevated SPAG5 expression and the expression of autophagy-related proteins, LC3-I, LC3-II, Beclin-1, and p62, in HUVECs. GFP-LC3 dots were increased in ox-LDL-treated HUVECs and LPS-treated HUVECs. SPAG5 knockdown reversed both ox-LDL and LPS treatment-mediated inhibition of cell proliferation and promotion of apoptosis in HUVECs. SPAG5 silencing further elevated autophagy and repressed the expression of PI3K, p-Akt/Akt, and p-mTOR/mTOR in ox-LDL-treated HUVECs. 3-MA (autophagy inhibitor) treatment reversed SPAG5 silencing-mediated increase of cell proliferation and decrease of apoptosis in ox-LDL-treated HUVECs. In vivo, SPAG5 knockdown reduced atherosclerotic plaques in AS mice through activating autophagy and inhibiting PI3K/Akt/mTOR signaling pathway. CONCLUSION: This work demonstrated that SPAG5 knockdown alleviated AS development through activating autophagy. Thus, SPAG5 may be a potential target for AS therapy.

IL-1ß Inhibition Partially Negates the Beneficial Effects of Diet-Induced Atherosclerosis Regression in Mice.

BACKGROUND: Thromboembolic events secondary to rupture or erosion of advanced atherosclerotic lesions is the global leading cause of death. The most common and effective means to reduce these major adverse cardiovascular events, including myocardial infarction and stroke, is aggressive lipid lowering via a combination of drugs and dietary modifications. However, we know little regarding the effects of reducing dietary lipids on the composition and stability of advanced atherosclerotic lesions, the mechanisms that regulate these processes, and what therapeutic approaches might augment the benefits of lipid lowering. METHODS: Smooth muscle cell lineage-tracing Apoe-/- mice were fed a high-cholesterol Western diet for 18 weeks and then a zero-cholesterol standard laboratory diet for 12 weeks before treating them with an IL (interleukin)-1ß or control antibody for 8 weeks. We assessed lesion size and remodeling indices, as well as the cellular composition of aortic and brachiocephalic artery lesions, indices of plaque stability, overall plaque burden, and phenotypic transitions of smooth muscle cell and other lesion cells by smooth muscle cell lineage tracing combined with single-cell RNA sequencing, cytometry by time-of-flight, and immunostaining plus high-resolution confocal microscopic z-stack analysis. RESULTS: Lipid lowering by switching Apoe-/- mice from a Western diet to a standard laboratory diet reduced LDL cholesterol levels by 70% and resulted in multiple beneficial effects including reduced overall aortic plaque burden, as well as reduced intraplaque hemorrhage and necrotic core area. However, contrary to expectations, IL-1ß antibody treatment after diet-induced reductions in lipids resulted in multiple detrimental changes including increased plaque burden and brachiocephalic artery lesion size, as well as increasedintraplaque hemorrhage, necrotic core area, and senescence as compared with IgG control antibody-treated mice. Furthermore, IL-1ß antibody treatment upregulated neutrophil degranulation pathways but downregulated smooth muscle cell extracellular matrix pathways likely important for the protective fibrous cap. CONCLUSIONS: Taken together, IL-1ß appears to be required for the maintenance of standard laboratory diet-induced reductions in plaque burden and increases in multiple indices of plaque stability.

Macrophage-Derived GSDMD Plays an Essential Role in Atherosclerosis and Cross Talk Between Macrophages via the Mitochondria-STING-IRF3/NF-κB Axis.

BACKGROUND: Macrophages play a crucial role in atherosclerotic plaque formation, and the death of macrophages is a vital factor in determining the fate of atherosclerosis. GSDMD (gasdermin D)-mediated pyroptosis is a programmed cell death, characterized by membrane pore formation and inflammatory factor release. METHODS: ApoE-/- and Gsdmd-/- ApoE-/- mice, bone marrow transplantation, and AAV (adeno-associated virus serotype 9)-F4/80-shGSDMD (shRNA-GSDMD) were used to examine the effect of macrophage-derived GSDMD on atherosclerosis. Single-cell RNA sequencing was used to investigate the changing profile of different cellular components and the cellular localization of GSDMD during atherosclerosis. RESULTS: First, we found that GSDMD is activated in human and mouse atherosclerotic plaques and Gsdmd-/- attenuates the atherosclerotic lesion area in high-fat diet-fed ApoE-/- mice. We performed single-cell RNA sequencing of ApoE-/- and Gsdmd-/- ApoE-/- mouse aortas and showed that GSDMD is principally expressed in atherosclerotic macrophages. Using bone marrow transplantation and AAV-F4/80-shGSDMD, we identified the potential role of macrophage-derived GSDMD in aortic pyroptosis and atherosclerotic injuries in vivo. Mechanistically, GSDMD contributes to mitochondrial perforation and mitochondrial DNA leakage and subsequently activates the STING (stimulator of interferon gene)-IRF3 (interferon regulatory factor 3)/NF-κB (nuclear factor kappa B) axis. Meanwhile, GSDMD regulates the STING pathway activation and macrophage migration via cytokine secretion. Inhibition of GSDMD with GSDMD-specific inhibitor GI-Y1 (GSDMD inhibitor Y1) can effectively alleviate the progression of atherosclerosis. CONCLUSIONS: Our study has provided a novel macrophage-derived GSDMD mechanism in the promotion of atherosclerosis and demonstrated that GSDMD can be a potential therapeutic target for atherosclerosis.

Interleukin-1 receptor accessory protein blockade limits the development of atherosclerosis and reduces plaque inflammation.

AIMS: The interleukin-1 receptor accessory protein (IL1RAP) is a co-receptor required for signalling through the IL-1, IL-33, and IL-36 receptors. Using a novel anti-IL1RAP-blocking antibody, we investigated the role of IL1RAP in atherosclerosis. METHODS AND RESULTS: Single-cell RNA sequencing data from human atherosclerotic plaques revealed the expression of IL1RAP and several IL1RAP-related cytokines and receptors, including IL1B and IL33. Histological analysis showed the presence of IL1RAP in both the plaque and adventitia, and flow cytometry of murine atherosclerotic aortas revealed IL1RAP expression on plaque leucocytes, including neutrophils and macrophages. High-cholesterol diet fed apolipoprotein E-deficient (Apoe-/-) mice were treated with a novel non-depleting IL1RAP-blocking antibody or isotype control for the last 6 weeks of diet. IL1RAP blockade in mice resulted in a 20% reduction in subvalvular plaque size and limited the accumulation of neutrophils and monocytes/macrophages in plaques and of T cells in adventitia, compared with control mice. Indicative of reduced plaque inflammation, the expression of several genes related to leucocyte recruitment, including Cxcl1 and Cxcl2, was reduced in brachiocephalic arteries of anti-IL1RAP-treated mice, and the expression of these chemokines in human plaques was mainly restricted to CD68+ myeloid cells. Furthermore, in vitro studies demonstrated that IL-1, IL-33, and IL-36 induced CXCL1 release from both macrophages and fibroblasts, which could be mitigated by IL1RAP blockade. CONCLUSION: Limiting IL1RAP-dependent cytokine signalling pathways in atherosclerotic mice reduces plaque burden and plaque inflammation, potentially by limiting plaque chemokine production.

Heparanase promotes the onset and progression of atherosclerosis in apolipoprotein E gene knockout mice.

BACKGROUND AND AIMS: Atherosclerosis is the primary underlying cause of myocardial infarction and stroke, which are the major causes of death globally. Heparanase (Hpse) is a pro-inflammatory extracellular matrix degrading enzyme that has been implicated in atherogenesis. However, to date the precise roles of Hpse in atherosclerosis and its mechanisms of action are not well defined. This study aims to provide new insights into the contribution of Hpse in different stages of atherosclerosis in vivo. METHODS: We generated Hpse gene-deficient mice on the atherosclerosis-prone apolipoprotein E gene knockout (ApoE-/-) background to investigate the impact of Hpse gene deficiency on the initiation and progression of atherosclerosis after 6 and 14 weeks high-fat diet feeding, respectively. Atherosclerotic lesion development, blood serum profiles, lesion composition and aortic immune cell populations were evaluated. RESULTS: Hpse-deficient mice exhibited significantly reduced atherosclerotic lesion burden in the aortic sinus and aorta at both time-points, independent of changes in plasma cholesterol levels. A significant reduction in the necrotic core size and an increase in smooth muscle cell content were also observed in advanced atherosclerotic plaques of Hpse-deficient mice. Additionally, Hpse deficiency reduced circulating and aortic levels of VCAM-1 at the initiation and progression stages of disease and circulating MCP-1 levels in the initiation but not progression stage. Moreover, the aortic levels of total leukocytes and dendritic cells in Hpse-deficient ApoE-/- mice were significantly decreased compared to control ApoE-/-mice at both disease stages. CONCLUSIONS: This study identifies Hpse as a key pro-inflammatory enzyme driving the initiation and progression of atherosclerosis and highlighting the potential of Hpse inhibitors as novel anti-inflammatory treatments for cardiovascular disease.

l-thyroxine attenuates extracellular Hsp90α-induced vascular endothelial calcification in diabetes mellitus, as revealed by parallel metabolic profiles.

BACKGROUND AND AIMS: Diabetic atherosclerotic vascular disease is characterized by extensive vascular calcification. However, an elevated blood glucose level alone does not explain this pathogenesis. We investigated the metabolic markers underlying diabetic atherosclerosis and whether extracellular Hsp90α (eHsp90α) triggers vascular endothelial calcification in this particular metabolic environment. METHODS: A parallel human/animal model metabolomics approach was used. We analyzed 40 serum samples collected from 24 patients with atherosclerosis and from the STZ-induced ApoE-/- mouse model. A multivariate statistical analysis of the data was performed, and mouse aortic tissue was collected for the assessment of plaque formation. In vitro, the effects of eHsp90α on endothelial cell calcification were assessed by serum analysis, Western blotting and immunoelectron microscopy. RESULTS: Diabetic ApoE-/- mice showed more severe plaque lesions and calcification damage. Stearamide, oleamide, l-thyroxine, l-homocitrulline and l-citrulline are biomarkers of diabetic ASVD; l-thyroxine was downregulated in both groups, and the thyroid sensitivity index was correlated with serum Hsp90α concentration. In vitro studies showed that eHsp90α increased Runx2 expression in endothelial cells through the LRP1 receptor. l-thyroxine reduced the increase in Runx2 levels caused by eHsp90α and affected the distribution and expression of LRP1 through hydrogen bonding with glutamine at position 1054 in the extracellular segment of LRP1. CONCLUSIONS: This study provides a mechanistic link between characteristic serum metabolites and diabetic atherosclerosis and thus offers new insight into the role of extracellular Hsp90α in promoting vascular calcification.

Resident Memory T Cells in the Atherosclerotic Lesion Associate With Reduced Macrophage Content and Increased Lesion Stability.

BACKGROUND: Tissue resident memory T (TRM) cells are a T-cell subset that resides at the site of prior antigen recognition to protect the body against reoccurring encounters. Besides their protective function, TRM cells have also been implicated in inflammatory disorders. TRM cells are characterized by the expression of CD69 and transcription factors Hobit (homolog of Blimp-1 [B lymphocyte-induced maturation protein 1] in T cells) and Blimp-1. As the majority of T cells in the arterial intima expresses CD69, TRM cells may contribute to the pathogenesis of atherosclerosis as well. Here, we aimed to assess the presence and potential role of TRM cells in atherosclerosis. METHODS: To identify TRM cells in human atherosclerotic lesions, a single-cell RNA-sequencing data set was interrogated, and T-cell phenotypes were compared with that of integrated predefined TRM cells. The presence and phenotype of TRM in atherosclerotic lesions was corroborated using a mouse model that enabled tracking of Hobit-expressing TRM cells. To explore the function of TRM cells during atherogenesis, RAG1-/- (recombination activating gene 1 deficient) LDLr-/- (low-density lipoprotein receptor knockout) mice received a bone marrow transplant from HobitKO/CREBlimp-1flox/flox mice, which exhibit abrogated TRM cell formation, whereafter the mice were fed a Western-type diet for 10 weeks. RESULTS: Human atherosclerotic lesions contained T cells that exhibited a TRM cell-associated gene signature. Moreover, a fraction of these T cells clustered together with predefined TRM cells upon integration. The presence of Hobit-expressing TRM cells in the atherosclerotic lesion was confirmed in mice. These lesion-derived TRM cells were characterized by the expression of CD69 and CD49α. Moreover, we demonstrated that this small T-cell subset significantly affects lesion composition, by reducing the amount of intralesional macrophages and increasing collagen content. CONCLUSIONS: TRM cells, characterized by the expression of CD69 and CD49α, constitute a minor population in atherosclerotic lesions and are associated with increased lesion stability in a Hobit and Blimp-1 knockout mouse model.

Immunoproteasomal Inhibition With ONX-0914 Attenuates Atherosclerosis and Reduces White Adipose Tissue Mass and Metabolic Syndrome in Mice.

BACKGROUND: Atherosclerosis is the major underlying pathology of cardiovascular disease and is driven by dyslipidemia and inflammation. Inhibition of the immunoproteasome, a proteasome variant that is predominantly expressed by immune cells and plays an important role in antigen presentation, has been shown to have immunosuppressive effects. METHODS: We assessed the effect of ONX-0914, an inhibitor of the immunoproteasomal catalytic subunits LMP7 (proteasome subunit ß5i/large multifunctional peptidase 7) and LMP2 (proteasome subunit ß1i/large multifunctional peptidase 2), on atherosclerosis and metabolism in LDLr-/- and APOE*3-Leiden.CETP mice. RESULTS: ONX-0914 treatment significantly reduced atherosclerosis, reduced dendritic cell and macrophage levels and their activation, as well as the levels of antigen-experienced T cells during early plaque formation, and Th1 cells in advanced atherosclerosis in young and aged mice in various immune compartments. Additionally, ONX-0914 treatment led to a strong reduction in white adipose tissue mass and adipocyte progenitors, which coincided with neutrophil and macrophage accumulation in white adipose tissue. ONX-0914 reduced intestinal triglyceride uptake and gastric emptying, likely contributing to the reduction in white adipose tissue mass, as ONX-0914 did not increase energy expenditure or reduce total food intake. Concomitant with the reduction in white adipose tissue mass upon ONX-0914 treatment, we observed improvements in markers of metabolic syndrome, including lowered plasma triglyceride levels, insulin levels, and fasting blood glucose. CONCLUSIONS: We propose that immunoproteasomal inhibition reduces 3 major causes underlying cardiovascular disease, dyslipidemia, metabolic syndrome, and inflammation and is a new target in drug development for atherosclerosis treatment.

Multifaceted roles of Meg3 in cellular senescence and atherosclerosis.

BACKGROUND AND AIMS: Long noncoding RNAs are involved in the pathogenesis of atherosclerosis. As long noncoding RNAs maternally expressed gene 3 (Meg3) prevents cellular senescence of hepatic vascular endothelium and obesity-induced insulin resistance, we decided to examine its role in cellular senescence and atherosclerosis. METHODS AND RESULTS: By analyzing our data and human and mouse data from the Gene Expression Omnibus database, we found that Meg3 expression was reduced in humans and mice with cardiovascular disease, indicating its potential role in atherosclerosis. In Ldlr-/- mice fed a Western diet for 12 weeks, Meg3 silencing by chemically modified antisense oligonucleotides attenuated the formation of atherosclerotic lesions by 34.9% and 20.1% in male and female mice, respectively, revealed by en-face Oil Red O staining, which did not correlate with changes in plasma lipid profiles. Real-time quantitative PCR analysis of cellular senescence markers p21 and p16 revealed that Meg3 deficiency aggravates hepatic cellular senescence but not cellular senescence at aortic roots. Human Meg3 transgenic mice were generated to examine the role of Meg3 gain-of-function in the development of atherosclerosis induced by PCSK9 overexpression. Meg3 overexpression promotes atherosclerotic lesion formation by 29.2% in Meg3 knock-in mice independent of its effects on lipid profiles. Meg3 overexpression inhibits hepatic cellular senescence, while it promotes aortic cellular senescence likely by impairing mitochondrial function and delaying cell cycle progression. CONCLUSIONS: Our data demonstrate that Meg3 promotes the formation of atherosclerotic lesions independent of its effects on plasma lipid profiles. In addition, Meg3 regulates cellular senescence in a tissue-specific manner during atherosclerosis. Thus, we demonstrated that Meg3 has multifaceted roles in cellular senescence and atherosclerosis.

Echocardiographic and pathologic identification of an aorto-left atrial fistula secondary to infective endocarditis in a canine patient.

A dog was presented for lameness, fever, and extreme lethargy. On physical exam, a new heart murmur, arrhythmia, and joint effusion were detected. These findings were not detected two months prior. A diagnostic work-up confirmed septic suppurative inflammation in multiple joints. Echocardiogram revealed aortic valvular endocarditis along with a communication, as a consequence of a fistula, that extended from just below the aortic sinotubular junction to the left atrial lumen. Due to a poor prognosis, humane euthanasia was elected. Necropsy and histopathology confirmed infective endocarditis of the aortic valve and an aorto-left atrial fistulous tract extending from the left coronary sinus of the aortic valve to the lumen of left atrium.

Deficiency of lncRNA MERRICAL abrogates macrophage chemotaxis and diabetes-associated atherosclerosis.

Diabetes-associated atherosclerosis involves excessive immune cell recruitment and plaque formation. However, the mechanisms remain poorly understood. Transcriptomic analysis of the aortic intima in Ldlr-/- mice on a high-fat, high-sucrose-containing (HFSC) diet identifies a macrophage-enriched nuclear long noncoding RNA (lncRNA), MERRICAL (macrophage-enriched lncRNA regulates inflammation, chemotaxis, and atherosclerosis). MERRICAL expression increases by 249% in intimal lesions during progression. lncRNA-mRNA pair genomic mapping reveals that MERRICAL positively correlates with the chemokines Ccl3 and Ccl4. MERRICAL-deficient macrophages exhibit lower Ccl3 and Ccl4 expression, chemotaxis, and inflammatory responses. Mechanistically, MERRICAL guides the WDR5-MLL1 complex to activate CCL3 and CCL4 transcription via H3K4me3 modification. MERRICAL deficiency in HFSC diet-fed Ldlr-/- mice reduces lesion formation by 74% in the aortic sinus and 86% in the descending aorta by inhibiting leukocyte recruitment into the aortic wall and pro-inflammatory responses. These findings unveil a regulatory mechanism whereby a macrophage-enriched lncRNA potently inhibits chemotactic responses, alleviating lesion progression in diabetes.

Paraspeckle protein NONO attenuates vascular calcification by inhibiting bone morphogenetic protein 2 transcription.

Vascular calcification is a pathological process commonly associated with atherosclerosis, chronic kidney disease, and diabetes. Paraspeckle protein NONO is a multifunctional RNA/DNA binding protein involved in many nuclear biological processes but its role in vascular calcification remains unclear. Here, we observed that NONO expression was decreased in calcified arteries of mice and patients with CKD. We generated smooth muscle-specific NONO-knockout mice and established three different mouse models of vascular calcification by means of 5/6 nephrectomy, adenine diet to induce chronic kidney failure, or vitamin D injection. The knockout mice were more susceptible to the development of vascular calcification relative to control mice, as verified by an increased calcification severity and calcium deposition. Likewise, aortic rings from knockout mice showed more significant vascular calcification than those from control mice ex vivo. In vitro, NONO deficiency aggravated high phosphate-induced vascular smooth muscle cell osteogenic differentiation and apoptosis, whereas NONO overexpression had a protective effect. Mechanistically, we demonstrated that the regulation of vascular calcification by NONO was mediated by bone morphogenetic protein 2 (BMP2). NONO directly bound to the BMP2 promoter using its C-terminal region, exerting an inhibitory effect on the transcription of BMP2. Thus, our study reveals that NONO is a novel negative regulator of vascular calcification, which inhibits osteogenic differentiation of vascular smooth muscle cell and vascular calcification via negatively regulating BMP2 transcription. Hence, NONO may provide a promising target for the prevention and treatment of vascular calcification.

Unraveling the Role of Sex in Endothelial Cell Dysfunction: Evidence From Lineage Tracing Mice and Cultured Cells.

BACKGROUND: Biological sex differences play a vital role in cardiovascular diseases, including atherosclerosis. The endothelium is a critical contributor to cardiovascular pathologies since endothelial cells (ECs) regulate vascular tone, redox balance, and inflammatory reactions. Although EC activation and dysfunction play an essential role in the early and late stages of atherosclerosis development, little is known about sex-dependent differences in EC. METHODS: We used human and mouse aortic EC as well as EC-lineage tracing (Cdh5-CreERT2 Rosa-YFP [yellow fluorescence protein]) atherosclerotic Apoe-/- mice to investigate the biological sexual dimorphism of the EC functions in vitro and in vivo. Bioinformatics analyses were performed on male and female mouse aortic EC and human lung and aortic EC. RESULTS: In vitro, female human and mouse aortic ECs showed more apoptosis and higher cellular reactive oxygen species levels than male EC. In addition, female mouse aortic EC had lower mitochondrial membrane potential (ΔΨm), lower TFAM (mitochondrial transcription factor A) levels, and decreased angiogenic potential (tube formation, cell viability, and proliferation) compared with male mouse aortic EC. In vivo, female mice had significantly higher lipid accumulation within the aortas, impaired glucose tolerance, and lower endothelial-mediated vasorelaxation than males. Using the EC-lineage tracing approach, we found that female lesions had significantly lower rates of intraplaque neovascularization and endothelial-to-mesenchymal transition within advanced atherosclerotic lesions but higher incidents of missing EC lumen coverage and higher levels of oxidative products and apoptosis. RNA-seq analyses revealed that both mouse and human female EC had higher expression of genes associated with inflammation and apoptosis and lower expression of genes related to angiogenesis and oxidative phosphorylation than male EC. CONCLUSIONS: Our study delineates critical sex-specific differences in EC relevant to proinflammatory, pro-oxidant, and angiogenic characteristics, which are entirely consistent with a vulnerable phenotype in females. Our results provide a biological basis for sex-specific proatherosclerotic mechanisms.

Working towards risk stratification for ascending aortic dilatation in pediatric Turner syndrome patients: results of a longitudinal echocardiographical observation.

This study aimed to longitudinally evaluate aortic root dimensions and elasticity in pediatric Turner syndrome (TS) in relation to known cardiac implications such as coarctation of the aorta (CoA) and bicuspid aortic valves (BAV) in order to create an improved risk profile for the presumed underlying vessel pathology in childhood. We report on the longitudinal findings of our pediatric TS outpatient clinic over a period of up to 7.6 years. Forty-nine TS patients (median age at baseline 9.7 ± 5.9 years, range 0-19.8) were followed-up for on average 2.9 ± 1.1 examinations and a median time of 3.4 ± 1.6 years. Aortic root (AoR) diameters and corresponding Z-scores were determined echocardiographically, and elasticity parameters as well as annual progression rates were calculated. At baseline, 16.3% of patients showed Z-scores > 2 at one or more levels of the AoR (35.7% of patients with BAV, odds ratio of 4.2). There was net progression to be noted at all measuring levels, leading to 28.6% of patients (50% of patients with BAV) exhibiting aortic dilatation at the end of follow-up. Progression correlated with the presence of BAV, non-mosaic monosomy, and age. A levelling-off of progression was seen with the onset of adolescence. CONCLUSIONS: Marked progression of aortic diameters leading to the development of dilatation can be observed in TS patients during childhood and stresses the importance of close surveillance during childhood. Main risk factors are BAV and complete monosomy 45X0. A beneficial influence of estrogen substitution can be suspected but needs further investigation. WHAT IS KNOWN: • Patients with Turner syndrome are at an increased risk for aortic dilatation and dissection. • The presence of BAV and complete monosomy 45X are additional risk factors. WHAT IS NEW: • Aortic dilatation can be detected in pediatric patients with Turner syndrome. • Relevant progression in childhood is possible in at-risk individuals and warrants close surveillance.

The activator protein-1 complex governs a vascular degenerative transcriptional programme in smooth muscle cells to trigger aortic dissection and rupture.

BACKGROUND AND AIMS: Stanford type A aortic dissection (AD) is a degenerative aortic remodelling disease marked by an exceedingly high mortality without effective pharmacologic therapies. Smooth muscle cells (SMCs) lining tunica media adopt a range of states, and their transformation from contractile to synthetic phenotypes fundamentally triggers AD. However, the underlying pathomechanisms governing this population shift and subsequent AD, particularly at distinct disease temporal stages, remain elusive. METHODS: Ascending aortas from nine patients undergoing ascending aorta replacement and five individuals undergoing heart transplantation were subjected to single-cell RNA sequencing. The pathogenic targets governing the phenotypic switch of SMCs were identified by trajectory inference, functional scoring, single-cell regulatory network inference and clustering, regulon, and interactome analyses and confirmed using human ascending aortas, primary SMCs, and a ß-aminopropionitrile monofumarate-induced AD model. RESULTS: The transcriptional profiles of 93 397 cells revealed a dynamic temporal-specific phenotypic transition and marked elevation of the activator protein-1 (AP-1) complex, actively enabling synthetic SMC expansion. Mechanistically, tumour necrosis factor signalling enhanced AP-1 transcriptional activity by dampening mitochondrial oxidative phosphorylation (OXPHOS). Targeting this axis with the OXPHOS enhancer coenzyme Q10 or AP-1-specific inhibitor T-5224 impedes phenotypic transition and aortic degeneration while improving survival by 42.88% (58.3%-83.3% for coenzyme Q10 treatment), 150.15% (33.3%-83.3% for 2-week T-5224), and 175.38% (33.3%-91.7% for 3-week T-5224) in the ß-aminopropionitrile monofumarate-induced AD model. CONCLUSIONS: This cross-sectional compendium of cellular atlas of human ascending aortas during AD progression provides previously unappreciated insights into a transcriptional programme permitting aortic degeneration, highlighting a translational proof of concept for an anti-remodelling intervention as an attractive strategy to manage temporal-specific AD by modulating the tumour necrosis factor-OXPHOS-AP-1 axis.

Histopathological Gap in Aortic Diseases: A Prospective Analysis.

Aortic dissection (AD) is a critical cardiovascular condition with the potential for devastating consequences. This study evaluated the histological changes in the aorta wall in patients with AD and aortic aneurysm (AA) who received surgical aortic replacement. Histopathological data showed that modifications of the media layer (p = 0.0197), myxomatous aspect (p = 0.0001), and subendothelial layer degeneration (p = 0.0107) were more frequently seen in AA versus AD samples. Patients with AA were approximately twice as likely to develop histological changes than those with AD (p = 0.0037). Patients with moderate or severe medial degeneration had a higher chance of developing AD (p = 0.0001). Because the histopathological score proved to be a predictor of both in-hospital and overall mortality, its evaluation should become the standard of care in any patients who undergo aortic replacement. Individualized postoperative management might be influenced by the histopathological aspect of the aortic layer.

Accelerated atherosclerosis in beta-thalassemia.

Children with beta-thalassemia (BT) present with an increase in carotid intima-medial thickness, an early sign suggestive of premature atherosclerosis. However, it is unknown if there is a direct relationship between BT and atherosclerotic disease. To evaluate this, wild-type (WT, littermates) and BT (Hbbth3/+) mice, both male and female, were placed on a 3-mo high-fat diet with low-density lipoprotein receptor suppression via overexpression of proprotein convertase subtilisin/kexin type 9 (PCSK9) gain-of-function mutation (D377Y). Mechanistically, we hypothesize that heme-mediated oxidative stress creates a proatherogenic environment in BT because BT is a hemolytic anemia that has increased free heme and exhausted hemopexin, heme's endogenous scavenger, in the vasculature. We evaluated the effect of hemopexin (HPX) therapy, mediated via an adeno-associated virus, to the progression of atherosclerosis in BT and a phenylhydrazine-induced model of intravascular hemolysis. In addition, we evaluated the effect of deferiprone (DFP)-mediated iron chelation in the progression of atherosclerosis in BT mice. Aortic en face and aortic root lesion area analysis revealed elevated plaque accumulation in both male and female BT mice compared with WT mice. Hemopexin therapy was able to decrease plaque accumulation in both BT mice and mice on our phenylhydrazine (PHZ)-induced model of hemolysis. DFP decreased atherosclerosis in BT mice but did not provide an additive benefit to HPX therapy. Our data demonstrate for the first time that the underlying pathophysiology of BT leads to accelerated atherosclerosis and shows that heme contributes to atherosclerotic plaque development in BT.NEW & NOTEWORTHY This work definitively shows for the first time that beta-thalassemia leads to accelerated atherosclerosis. We demonstrated that intravascular hemolysis is a prominent feature in beta-thalassemia and the resulting increases in free heme are mechanistically relevant. Adeno-associated virus (AAV)-hemopexin therapy led to decreased free heme and atherosclerotic plaque area in both beta-thalassemia and phenylhydrazine-treated mice. Deferiprone-mediated iron chelation led to deceased plaque accumulation in beta-thalassemia mice but provided no additive benefit to hemopexin therapy.

The Genetic Determinants of Aortic Distention.

BACKGROUND: As the largest conduit vessel, the aorta is responsible for the conversion of phasic systolic inflow from ventricular ejection into more continuous peripheral blood delivery. Systolic distention and diastolic recoil conserve energy and are enabled by the specialized composition of the aortic extracellular matrix. Aortic distensibility decreases with age and vascular disease. OBJECTIVES: In this study, we sought to discover epidemiologic correlates and genetic determinants of aortic distensibility and strain. METHODS: We trained a deep learning model to quantify thoracic aortic area throughout the cardiac cycle from cardiac magnetic resonance images and calculated aortic distensibility and strain in 42,342 UK Biobank participants. RESULTS: Descending aortic distensibility was inversely associated with future incidence of cardiovascular diseases, such as stroke (HR: 0.59 per SD; P = 0.00031). The heritabilities of aortic distensibility and strain were 22% to 25% and 30% to 33%, respectively. Common variant analyses identified 12 and 26 loci for ascending and 11 and 21 loci for descending aortic distensibility and strain, respectively. Of the newly identified loci, 22 were not significantly associated with thoracic aortic diameter. Nearby genes were involved in elastogenesis and atherosclerosis. Aortic strain and distensibility polygenic scores had modest effect sizes for predicting cardiovascular outcomes (delaying or accelerating disease onset by 2%-18% per SD change in scores) and remained statistically significant predictors after accounting for aortic diameter polygenic scores. CONCLUSIONS: Genetic determinants of aortic function influence risk for stroke and coronary artery disease and may lead to novel targets for medical intervention.

Number of calcified aortic valve leaflets: natural history and prognostic value in patients undergoing haemodialysis.

AIMS: Aortic valve calcification in aortic sclerosis, a precursor of aortic stenosis (AS), is not always present in all three leaflets; how calcification develops in each leaflet is unknown. We aimed to investigate the natural history of calcification development in each aortic valve leaflet and the prognostic value of the number of calcified leaflets. METHODS AND RESULTS: In a retrospective multicentre cohort study of patients undergoing haemodialysis without AS, we observed calcification development in each aortic valve leaflet using echocardiography. We investigated the association between the number of calcified leaflets and AS development and mortality using time-to-event analysis. Among the 1507 patients (mean age, 66 years; 66% male) included in the longitudinal echocardiography analysis, 709 (47%) had aortic sclerosis at baseline: one-leaflet calcified, 370 (52%); two-leaflet calcified, 215 (30%); and three-leaflet calcified, 124 (17%). The median time for one calcified leaflet increase was 3-4 years, and 251 (17%) patients developed AS during a median 3.2-year follow-up. The increased number of calcified aortic valve leaflets was associated with developing AS; compared with that of one-leaflet calcified, the adjusted hazard ratios (aHRs) [95% confidence intervals (CIs)] of two- and three-leaflet calcified were 2.12 (1.49-3.00) and 4.43 (3.01-6.52), respectively; the aHR (95% CI) per one calcified leaflet increase was 2.24 (1.96-2.55). It was also associated with all-cause mortality; the aHR (95% CI) per one calcified leaflet increase was 1.18 (1.08-1.27). CONCLUSION: The number of calcified aortic valve leaflets strongly predicted AS development and even mortality in patients undergoing haemodialysis, suggesting the usefulness of assessing calcification for each valve leaflet separately using echocardiography.

A long-term survival rat model of spinal cord ischemia injury: Thoracic aortic occlusion combined with aortic bypass circulation.

OBJECTIVE: This study aims to investigate the methods for rat spinal cord ischemia injury models with a high long-term survival rate. METHODS: The rats were divided into three groups: the treatment group, the control group, and the sham operation group. The treatment group had a blocked thoracic aorta (landing zone 3 by Ishimaru - T11) + aortic bypass circulation for 20 min. In the control group, the thoracic aorta at the landing zone 3 was blocked for 20 min. In the sham operation group, only thoracotomy without thoracic aortic occlusion was performed. The mean arterial blood pressure (MABP) of the thoracic aorta and caudal artery before and after thoracic aortic occlusion was monitored intraoperatively. Spinal cord function was monitored by a transcranial motor evoked potential (Tc-MEP) during the operation. Spinal cord function was evaluated by the BBB scale (Basso, Beattie, & Bresnahan locomotor rating scale) scores at multiple postoperative time points. The spinal cord sections of the rats were observed for 7 days after surgery, and the survival curves were analyzed for 28 days after surgery. RESULTS: After aortic occlusion, the MABP of thoracic aorta decreased to 6% of that before occlusion, and the MABP of caudal artery decreased to 63% of that before occlusion in the treatment group. In the control group, the MABP of both thoracic aorta and caudal artery decreased to 19% of that before occlusion. The Tc-MEP waveform of the treatment group disappeared after 6 min, and that of the control group disappeared after 8 min until the end of surgery. There was no change in the Tc-MEP waveform in the sham operation group. The BBB score of the treatment group decreased more obviously than the control group, and there was a significant difference. There was no decrease in the sham group. Spinal cord sections showed a large number of degeneration and necrosis of neurons, infiltration of inflammatory cells, and proliferation of surrounding glial cells in the treatment group. In the control group, multiple neurons were necrotic. The histology of the sham operation group was normal. The 28-day survival rate of the treatment group was 73.3%, which was higher than the control group (40.0%), and there was a significant difference (p < 0.05). CONCLUSION: Thoracic aortic occlusion combined with aortic bypass is an effective modeling method for rats with accurate modeling effects and high long-term survival rates.