BACKGROUND: Ischemic stroke requires effective reperfusion therapies to limit brain injury, yet rtPA (recombinant tissue-type plasminogen activator) efficacy is limited, particularly in platelet-rich thrombi. Neutrophil...BACKGROUND: Ischemic stroke requires effective reperfusion therapies to limit brain injury, yet rtPA (recombinant tissue-type plasminogen activator) efficacy is limited, particularly in platelet-rich thrombi. Neutrophil extracellular traps (NETs) and their components, especially histones and DNA, contribute to thrombolysis resistance. Chondroitin sulfate (CS), a glycosaminoglycan with high affinity for extracellular histones, may neutralize their prothrombotic effects and improve outcomes. This study aimed to evaluate the effects of CS in preclinical ischemic stroke models and its impact on components of neutrophil extracellular traps. METHODS: Two mouse models of middle cerebral artery occlusion were used: a fibrin-rich thromboembolic stroke model (rtPA-sensitive) and a platelet-rich aluminum chloride model (rtPA-resistant). Mice received intravenous CS (30-120 mg/kg), rtPA (10 mg/kg), or a combination of both. Lesion volume, tissue recanalization/reperfusion, hemorrhagic transformation, and functional connectivity were assessed via 7T magnetic resonance imaging and ultrafast Doppler imaging. In vitro coagulation-fibrinolysis assays examined the effects of neutrophil extracellular trap components on fibrin polymerization and fibrinolysis, and their modulation by CS±rtPA. RESULTS: In the fibrin-rich model, CS alone reduced lesion volume by 36% and improved recanalization, comparable to rtPA (43%), without increasing hemorrhagic transformation. CS enhanced functional connectivity recovery at 24 hours, whereas combined CS+rtPA lost these benefits. In the platelet-rich model, CS did not affect lesion size, recanalization, or hemorrhage. In vitro, histones promoted clot stabilization and altered fibrinolysis, effects fully neutralized by equimolar CS in the absence of rtPA. With rtPA, CS's neutralizing capacity was reduced, and histone-driven profibrinolysis was accentuated at higher CS doses. DNA produced opposite effects to histones, and combined DNA+histones masked histone activity, resisting inhibition by CS+DNase. CONCLUSIONS: CS mitigates histone-mediated prothrombotic effects, improves reperfusion and network recovery in fibrin-rich stroke, but loses efficacy in platelet-rich thrombi and when combined with rtPA. These findings support CS as a potential adjunct or alternative therapy, particularly for patients with contraindications to rtPA.
Zhong L, Sun Y, Chen G
… +11 more, Wang J, Yang Z, Lu W, Xiao X, Song H, Xiong J, Lin B, Wang K, Yang S, Bin J, Jiang X
Arterioscler Thromb Vasc Biol
· 2025 Dec · PMID 41164881
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BACKGROUND: Abdominal aortic aneurysm (AAA), a pathological dilation of the abdominal aorta, is primarily driven by chronic aortic wall inflammation. The well-established anti-inflammatory microRNA 146a (miR-146a) has be...BACKGROUND: Abdominal aortic aneurysm (AAA), a pathological dilation of the abdominal aorta, is primarily driven by chronic aortic wall inflammation. The well-established anti-inflammatory microRNA 146a (miR-146a) has been implicated as a key regulator in various chronic inflammatory pathologies. However, its potential functional role in the pathogenesis of AAA remains to be elucidated. METHODS: We constructed Ang II (angiotensin II)-induced and PPE (porcine pancreatic elastase)-induced models in global miR-146a knockout mice, vascular smooth muscle cell (VSMC)-specific miR-146a knockout mice, and macrophage-specific miR-146a knockout mice, respectively, to explore the role of miR-146a in AAA. Western blot, quantitative polymerase chain reaction, and immunohistochemistry were used to detect the levels of aortic proinflammatory markers and VSMC contractile proteins, whereas flow cytometry was used to assess M1/M2-like macrophage polarization. To validate the downstream mechanism, dibenzazepine was intraperitoneally injected to inhibit the Notch1 pathway in rescue experiments. RESULTS: In the Ang II-induced and PPE-induced model, global knockout of miR-146a promoted AAA development, increased maximal aortic diameter, exacerbated medial elastin degradation, and upregulated aortic proinflammatory markers (COX2 [cyclooxygenase 2], MMP [matrix metalloproteinase] 2, MMP9, and CCL2 [chemokine (C-C motif) ligand 2]). Flow cytometry analysis revealed that global miR-146a deficiency also induced macrophage polarization toward the inflammatory M1 phenotype. Conditional deletion of miR-146a in VSMCs and macrophages largely replicated AAA formation and proinflammatory effects. Furthermore, AAV9 (adeno-associated virus)-mediated miR-146a knockdown significantly reduced VSMC contractile proteins CNN1 (calponin 1), SM22α (smooth muscle 22α), and α-SMA (α-smooth muscle actin) in mouse aortas at 7 days post-Ang II perfusion. Mechanistically, Notch1 antagonist dibenzazepine effectively rescued AAA characteristics and M1 biomarkers while enhancing M2 biomarkers in global miR-146a knockout mice. CONCLUSIONS: The absence of miR-146a potentiates AAA formation by promoting VSMC phenotypic switching, Notch1 signaling-mediated aortic inflammation, and macrophage M1 polarization. Thus, miR-146a plays a critical role in maintaining aortic structural integrity to prevent aneurysmal pathogenesis.
Tian W, Singh K, Kajuluri LP
… +27 more, Lu J, Lee S, Roh K, Nicholson CJ, Jiang W, Barnes HJ, Sato T, Boerboom S, Ostrom K, Li RH, Birchenough C, Moore E, Tattersfield H, Sigurslid HH, Mahamdeh MS, Mosquera JV, Hodonsky CJ, Johnson AL, Chou EL, Nigwekar S, Lindsay ME, Bloch DB, Lino Cardenas CL, Ichinose F, Miller CL, Wein MN, Malhotra R
Arterioscler Thromb Vasc Biol
· 2025 Dec · PMID 41164880
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BACKGROUND: Vascular calcification is an active process driven by osteogenic phenotypic transition of vascular smooth muscle cells (VSMCs) and regulated by a bone-related gene regulatory network. Recent studies showed th...BACKGROUND: Vascular calcification is an active process driven by osteogenic phenotypic transition of vascular smooth muscle cells (VSMCs) and regulated by a bone-related gene regulatory network. Recent studies showed that FAK (focal adhesion kinase) regulates bone formation by affecting the cellular localization of HDAC (histone deacetylase) 4 and HDAC5. However, it is not known whether FAK exerts effects on vascular calcification in VSMCs through regulating HDACs. METHODS: We used perturbational assays to assess the role of HDAC4, HDAC5, and FAK in VSMC calcification. Pharmacological inhibition and gene silencing of FAK were used to evaluate effects on calcification, cell migration, and the expression of procalcification factors. Leptomycin was used to inhibit the nuclear export of HDACs. In addition, ex vivo cultures of mouse and human arteries were treated with a FAK inhibitor to assess effects on arterial calcification. Single-cell transcriptomic expression of FAK was examined in healthy and diseased human coronary arteries. RESULTS: HDAC4 and HDAC5 were identified as positive regulators of vascular calcification. Pharmacological inhibition or gene silencing of FAK blocked VSMC calcification, abrogated the osteogenic medium-induced elevation of procalcification factors, and reduced cell migration. FAK inhibition reduced HDAC4 and HDAC5 phosphorylation and enhanced nuclear localization of these HDAC proteins. Inhibition of HDAC4 and HDAC5 nuclear export with leptomycin showed similar effects on calcification as FAK inhibition. Treatment with the FAK inhibitor attenuated the calcification of ex vivo mouse and human arteries. FAK gene expression was dysregulated in human diseased coronary arteries compared with healthy coronary arteries, and, in single-cell analysis of human arterial tissue, FAK expression was highest in VSMCs at an intermediate state between contractile and osteogenic phenotypes. CONCLUSIONS: FAK promotes VSMC calcification, at least in part, via phosphorylation of HDAC4 and HDAC5. Targeted regulation of the activity of FAK, HDAC4, and HDAC5 may be an effective strategy for the treatment of vascular calcification.
Johnson CF, Schafer CM, Burge KY
… +3 more, Coon BG, Chaaban H, Griffin CT
Arterioscler Thromb Vasc Biol
· 2026 Jan · PMID 41164879
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BACKGROUND: RIPK3 (receptor-interacting protein kinase 3) has context-specific roles that are frequently associated with cellular damage and death. We previously found that hypoxia can trigger elevated levels of RIPK3 in...BACKGROUND: RIPK3 (receptor-interacting protein kinase 3) has context-specific roles that are frequently associated with cellular damage and death. We previously found that hypoxia can trigger elevated levels of RIPK3 in endothelial cells (ECs), which contributes to lethal vascular rupture during mouse embryonic development. However, it is unknown whether elevated RIPK3 likewise compromises endothelial barrier function in adult vasculature under hypoxic conditions such as ischemia-reperfusion (I/R) injury. METHODS: Twelve-week-old male and female littermate control or inducible EC-specific knockout () mice were exposed to surgical intestinal I/R injury. Clodronate liposomes were used to reduce circulating monocytes in vivo. Immortalized murine EC (MS1 [mile sven 1) and murine macrophage (BMA3.1A7 [bone marrow A clone 3.187]) lines were used for in vitro experiments. RESULTS: mice displayed an unexpected increase in small intestinal vascular permeability after I/R injury, rather than the decrease we predicted. Subsequent analyses using multiplex cytokine assays revealed significantly elevated levels of IL-6 (interleukin-6) in the serum and small intestinal tissue of I/R-injured mice. Upon TNFα (tumor necrosis factor-alpha) stimulation of immortalized murine knockout ECs grown in vitro, we found increased transcription and secretion of IL-6. These cells also expressed elevated levels of VCAM-1 (vascular cell adhesion molecule-1), which was likewise upregulated in the small intestines of mice. Using an IL-6 neutralizing antibody, we found that IL-6 triggered VCAM-1 elevation in knockout cells. This VCAM-1 expression correlated with enhanced macrophage binding to knockout cells and increased accumulation of leukocytes in small intestines following I/R injury. Importantly, reduction of circulating monocytes with clodronate liposomes led to rescue of IR injury-induced vascular permeability in mice. CONCLUSIONS: Endothelial RIPK3 suppresses EC activation and inflammation associated with IL-6 and VCAM-1 elevation to protect the vascular barrier in the context of intestinal I/R injury. Thus, endothelial RIPK3 plays surprisingly beneficial roles that reduce I/R injury-induced vascular dysfunction.
Schrammel A, Wölkart G, Ableitner E
… +13 more, Derler M, Potoschnig I, Schoiswohl G, Haemmerle G, Wolfrum C, Kershaw EE, Schmid ST, Abdellatif M, Sedej S, Zechner R, Schweiger M, Mayer B, Mussbacher M
Arterioscler Thromb Vasc Biol
· 2025 Dec · PMID 41164878
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BACKGROUND: Perivascular adipose tissue (PVAT) fine-tunes blood vessel contractility and vascular homeostasis. During obesity and atherosclerosis, PVAT becomes dysfunctional and loses its anticontractile potential. Previ...BACKGROUND: Perivascular adipose tissue (PVAT) fine-tunes blood vessel contractility and vascular homeostasis. During obesity and atherosclerosis, PVAT becomes dysfunctional and loses its anticontractile potential. Previously, we reported that global knockout of adipose triglyceride lipase (ATGL), the major enzyme responsible for the breakdown of triglycerides, has the potential to modify PVAT functions. To address the causal relationship between PVAT lipolysis and blood vessel contractility, we analyzed ex vivo vasomotor function of mice with tissue-specific rescue/overexpression or knockout of ATGL in adipose tissue. METHODS: To generate mice lacking ATGL in all tissues except for adipose tissue (ATGL knockout with adipocyte-specific expression of ATGL [A+/AKO]), we crossed adipocyte ATGL-rescued (A+) mice with ATGL-deficient (ATGL knockout [AKO]) mice. Body weight, plasma levels of fatty acids, and blood glucose were compared between A+/AKO and AKO mice. Ex vivo vasoreactivity studies were performed in the absence and presence of PVAT to test for acute and chronic effects of PVAT on vascular function. RESULTS: Adipocyte-rescued AKO mice (A+/AKO) had significantly less amounts of PVAT than AKO controls while displaying moderate ATGL expression. A+/AKO aortas exhibited decreased anticontractile effects of PVAT compared with AKO aortas. This effect on contractile function was observed in an agonist-specific manner without affecting smooth muscle cell function or endothelium-dependent relaxation. Assessment of cardiac function using the Langendorff setup revealed that adipocyte ATGL selectively modulated vascular contractility without affecting systolic or diastolic performance. Studies using mice that express ATGL solely in cardiac muscle and adipocyte-specific ATGL knockout mice verified our findings in A+/AKO mice, revealing acute and chronic effects of adipocyte lipolysis on vasoreactivity. CONCLUSIONS: We provide the first evidence that changes in adipocyte lipolysis have the potential to regulate blood vessel contractility. Ablation of ATGL in adipocytes decreases vascular contractility and, thus, has the potential to prevent PVAT dysfunction in obesity and atherosclerosis.
Polkinghorne MD, Badi I, Baragetti A
… +18 more, Chauhan J, Xie C, Wahome E, Akoumianakis I, Foran D, Patel P, de Araujo E, Kotanidis CP, Krasopoulos G, Sayeed R, Srivastava V, Kourliouros A, Walcot N, Sastry P, Guzik T, Channon KM, Norata GD, Antoniades C
Arterioscler Thromb Vasc Biol
· 2026 Jan · PMID 41164877
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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...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 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) (=0.03) and LPA GRS (=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 (=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]; =0.82) or oxidative stress (ie, malondialdehyde; =0.61). Finally, both LPA GRS (hazard ratio, 3.615 [95% CI, 1.044-12.515]; =0.043) and high plasma Lp(a) (hazard ratio, 3.286 [95% CI, 1.003-10.767]; =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: The alternative pathway (AP) plays a crucial role in triggering complement activation and promoting chronic inflammation. This study aims to investigate the longitudinal association between AP and atheroscler...BACKGROUND: The alternative pathway (AP) plays a crucial role in triggering complement activation and promoting chronic inflammation. This study aims to investigate the longitudinal association between AP and atherosclerosis, and explore the potential role of gut microbiota and inflammatory factors in their association. METHOD: This study was based on a 9-year prospective cohort of 3382 participants from Guangzhou, China (mean age±SD, 57.75±5.85 years; 68.8% female), with data on serum APACPs (AP-associated complement proteins) and carotid plaque (measured by ultrasound) repeatedly measured up to 3×. Baseline inflammatory markers were evaluated in 923 participants, and gut shotgun metagenome data were obtained from 1567 participants. Mendelian randomization analysis was performed using genome-wide significant genetic variants as instrumental variables to suggest potential causal associations. RESULTS: Both longitudinal and prospective analyses consistently demonstrated positive associations between carotid plaque and 3 complement components: C3 (complement C3; odds ratios [95% Cl] for the highest versus lowest quartiles, 1.36 [1.07-1.74] in longitudinal analysis and 1.29 [1.06-1.56] in prospective analysis), CFB (complement factor B; 1.36 [1.07-1.72] in longitudinal analysis and 1.39 [1.15-1.69] in prospective analysis), and CFH (complement factor H; 1.39 [1.10-1.76] in longitudinal analysis and 1.31 [1.07-1.61] in prospective analysis). Mendelian randomization analysis suggested a potential causal association between CFB and carotid plaque. Inflammatory factors (CRP [C-reactive protein] and IL-6 [interleukin-6]) and microbial species (, , , , , and ) were significantly associated with both APACPs and carotid plaque (<0.05). For example, butyrate-producing bacterium was inversely associated with CFB and carotid plaque (odds ratios [95% CI], 0.83 [0.79-0.88]) and may mediate the CFB-carotid plaque association (proportion mediated, 13.5%; =0.005). Microbial risk score (weighted sum of selected microbial species; proportion mediated, 42.6%; <0.001) and total immune factors (the sum of all inflammatory factors; proportion mediated, 19.0%; =0.002) mediated the association between Total-APACPs (sum of standardized carotid plaque-related APACPs [C3, CFB, and CFH]) and carotid plaque. CONCLUSIONS: Our study showed a negative association between the AP and carotid plaque in a longitudinal cohort. Gut microbiota and inflammatory biomarkers may provide mechanistic insights into the association between the AP and atherosclerosis. Our findings pave the way for the development of new therapeutic targets for atherosclerosis.
Schrenk S, Sherpa C, Bischoff LJ
… +2 more, Cai Y, Boscolo E
Arterioscler Thromb Vasc Biol
· 2025 Dec · PMID 41127911
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BACKGROUND: Venous malformations (VMs) are developmental defects of the vasculature characterized by tremendously enlarged and dysfunctional veins. Gain-of-function somatic mutations in TIE2 (endothelial tyrosine kinase...BACKGROUND: Venous malformations (VMs) are developmental defects of the vasculature characterized by tremendously enlarged and dysfunctional veins. Gain-of-function somatic mutations in TIE2 (endothelial tyrosine kinase receptor) have been identified as the leading driver of VM pathogenesis. The aim of this study was to determine whether the aberrant venous lumen expansion is caused by recruitment of wild-type (WT) endothelial cells (EC) to the lesion or by TIE2-mutant EC clonal expansion. METHODS: To investigate the contribution of TIE2-mutant EC and WT EC to the aberrant venous lumen expansion, we used a xenograft murine model of VM generated with a combination of TIE2-mutant EC and WT EC. To perform longitudinal studies, we used a 3-dimensional fibrin gel lumen formation assay and a migration assay, both using WT EC in competition or confrontation with TIE2-mutant EC. To investigate the mechanisms implicated in VM lumen expansion, we used RNA-sequencing and short-hairpin RNA silencing in the TIE2-mutant EC. RESULTS: We demonstrate here that in the VM xenograft model, the aberrant blood vessels were lined almost exclusively by TIE2-mutant EC, and WT EC were rarely found. Functionally, the TIE2-mutant EC exerted a competitive advantage over WT EC by inhibiting WT EC sprouting. In line with these findings, TIE2-mutant EC promoted repulsion of WT EC. Short-hairpin RNA-mediated silencing of Sema (Semaphorin) 3A or Sema3F in TIE2-mutant EC rescued this chemorepellent phenotype and restored the ability of WT EC to migrate, sprout, and form lumens. Furthermore, knockdown of Sema3A or Sema3F in TIE2-mutant EC normalized the blood vessel size in vivo. CONCLUSIONS: Our results demonstrate that WT EC are not recruited to the aberrant veins, suggesting that VM pathogenesis is fueled by clonal expansion of TIE2-mutant EC. Mechanistically, we show that Sema3A and Sema3F are overexpressed in TIE2-mutant EC and play a crucial role in the pathological vascular lumen expansion in VM.
Arterioscler Thromb Vasc Biol
· 2025 Dec · PMID 41127910
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BACKGROUND: Pericytes are vascular mural cells, critical to the formation and maintenance of tightly regulated microvascular networks, including the inner blood-retinal barrier, and their dysfunction is characteristic of...BACKGROUND: Pericytes are vascular mural cells, critical to the formation and maintenance of tightly regulated microvascular networks, including the inner blood-retinal barrier, and their dysfunction is characteristic of many vascular diseases, including diabetic retinopathy. Although donor eye and animal studies have suggested a link between pericyte loss and retinal vascular leakage, this relationship has not been explored in living humans, nor has the role of pericytes in predicting other complications, like albuminuria, in diabetes. METHODS: In this pilot study, we utilized adaptive optics scanning laser ophthalmoscopy to image and quantify retinal capillary pericytes in humans with diabetes. Leakage was manually delineated on fluorescein angiography to calculate macular leakage (%), whereas optical coherence tomography and optical coherence tomography-angiography evaluated retinal thickness and capillary nonperfusion, respectively. In addition, urine albumin/creatinine ratios (mg/g) were extracted from charts, post hoc, to evaluate the relationship between retinal pericytes and albuminuria in a subpopulation (n=14). RESULTS: The study included 24 eyes from 23 patients with a range of diabetic retinopathy severity spanning from no diabetic retinopathy to proliferative diabetic retinopathy. Notably, capillary pericyte density showed a strong, negative correlation to macular leakage (=-0.68; <0.001) and moderate correlation to optical coherence tomography thickness (=-0.45; =0.027). Receiver operating characteristics analysis identified pericyte density thresholds (≤13.2 and 13.4 pericytes per 100 µm) that showed high area under the curves for detecting macular leakage (area under the curve, 0.87) and volumetric thickness (area under the curve, 0.80). Interestingly, albumin/creatinine ratios were significantly higher in individuals with ≤13.2 retinal capillary pericyte per 100 µm compared with those above the threshold (n=14, =0.046). CONCLUSIONS: Retinal capillary pericyte density, quantified in vivo, correlates significantly with macular thickening and angiographic leakage. We identified a threshold of pericyte loss that distinguishes individuals based on macular leakage status and albuminuria, providing important insights into retinal pericytes and their potential utility as a biomarker of vascular permeability.
Zhou X, Pucel JC, Nomura-Kitabayashi A
… +10 more, Chandakkar P, Guidroz AP, Jhangiani NL, Bao D, Fan J, Arthur HM, Ullmer C, Klein C, Marambaud P, Meadows SM
Tilp A, Nasias D, Carley AL
… +10 more, Park MY, Mooring A, Tirumalasetty MB, Abumrad NA, Wang Y, Miao QR, Lewandowski ED, Alemán JO, Goldberg IJ, Cabodevilla AG
Arterioscler Thromb Vasc Biol
· 2025 Dec · PMID 41099102
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BACKGROUND: Movement of circulating lipids into tissues and arteries requires transfer across the endothelial cell (EC) barrier. This process allows the heart to obtain fatty acids, its chief source of energy, and apoB-c...BACKGROUND: Movement of circulating lipids into tissues and arteries requires transfer across the endothelial cell (EC) barrier. This process allows the heart to obtain fatty acids, its chief source of energy, and apoB-containing lipoproteins to cross the arterial endothelial barrier, leading to cholesterol accumulation in the subendothelial space. Multiple studies have established elevated postprandial TRLs (triglyceride-rich lipoproteins) as an independent risk factor for cardiovascular disease. We explored how chylomicrons affect ECs and transfer their fatty acids across the EC barrier. METHODS: We had reported that media from chylomicron-treated ECs lead to lipid droplet formation in macrophages. To determine the responsible component of this media, we assessed whether removing the extracellular vesicles (EVs) would obviate this effect. EVs from control and treated cells were then characterized by protein, lipid, and microRNA content. We also studied the EV-induced transcription changes in macrophages and ECs and whether knockdown of SR-BI (scavenger receptor-BI) altered these responses. In addition, using chylomicrons labeled with [C]oleate, we studied the uptake and release of this labeled by ECs. RESULTS: Chylomicron treatment of ECs led to an inflammatory response that included production of EVs that drove macrophage lipid droplet accumulation. The EVs contained little free fatty acids and triglycerides, but abundant phospholipids and diacylglycerols. In concert with this, []C labeled chylomicron triglycerides exited ECs primarily in phospholipids. EVs from chylomicron-treated versus untreated ECs were larger, more abundant, and contained specific microRNAs. Treatment of macrophages and naive ECs with media from chylomicron-treated ECs increased expression of inflammatory genes. CONCLUSIONS: EC chylomicron metabolism produces EVs that increase macrophage inflammation and create LDs. Media containing these EVs also increases EC inflammation, illustrating an autocrine inflammatory process. Fatty acids within chylomicron triglycerides are converted to phospholipids within EVs. Thus, EC uptake of chylomicrons constitutes an important pathway for vascular inflammation and tissue lipid acquisition.
D'Erasmo L, Tramontano D, Di Costanzo A
… +11 more, Casula M, Galimberti F, Baratta F, Cefalù AB, Tarugi PM, Calandra S, Zambon A, Averna M, Catapano AL, Arca M, LIPIGEN-sHTG Group
Arterioscler Thromb Vasc Biol
· 2025 Dec · PMID 41099101
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BACKGROUND: We aimed to compare the molecular and clinical characteristics of patients identified in Italy as affected by either familial chylomicronemia syndrome (FCS) or multifactorial chylomicronemia syndrome (MCS) an...BACKGROUND: We aimed to compare the molecular and clinical characteristics of patients identified in Italy as affected by either familial chylomicronemia syndrome (FCS) or multifactorial chylomicronemia syndrome (MCS) and to assess the overall benefit of novel triglyceride-lowering therapies prescribed to these patients within the routine clinical care. METHODS: From the national LIPIGEN-sHTG (Lipid Transport Disorders Italian Genetic Network-Severe Hypertriglyceridemia) registry, 169 patients (57 FCS, 51 MCS, 61 variant-negative, variant-negative MCS) were retrospectively analyzed. Data on clinical and genetic characteristics, medical history, and medications were collected. Peak triglyceride levels were used to define untreated lipid phenotypes. RESULTS: In FCS, 72% exhibited biallelic and 28% variants; in MCS, 38% (n=19) carried variants, and 38% (n=19) carried variants, whereas the remaining individuals were carriers of (n=3), (n=2), and or variants (n=8), respectively. Peak TGs were highest in FCS (3000 mg/dL [interquartile range, 2116.0-4265.0]), followed by MCS (1817 mg/dL [interquartile range, 1370.0-3062.0]) and variant-negative MCS (1340.0 mg/dL [interquartile range, 946.5-2508.5]; <0.001). FCS showed a 3.4-fold higher risk of acute pancreatitis than others, whereas no significant differences were observed between groups in the prevalence of atherosclerotic cardiovascular diseases. In the subset of patients with FCS receiving novel therapies (lomitapide or volanesorsen; 35%), triglyceride levels decreased by 62%, as compared with an 11% reduction in those on conventional treatment. Across the cohort, posttreatment triglyceride levels were 895 mg/dL in FCS, 352 mg/dL in MCS, and 386 mg/dL in variant-negative MCS. CONCLUSIONS: As compared with MCS, patients with FCS showed a more severe phenotype and higher prevalence of variants. Lomitapide and volanesorsen provide better triglyceride control, yet only one-third of FCS were treated with these drugs in the routine clinical practice.
Arterioscler Thromb Vasc Biol
· 2025 Dec · PMID 41064860
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The lung lymphatics are formed from lymphatic endothelial cells and play a pivotal role in lung fluid homeostasis and immune trafficking. Though blood vascular function in the lung has long been an area of active investi...The lung lymphatics are formed from lymphatic endothelial cells and play a pivotal role in lung fluid homeostasis and immune trafficking. Though blood vascular function in the lung has long been an area of active investigation, many aspects of lung lymphatic vascular function have only recently been uncovered. In this review, we will discuss our current knowledge of lung lymphatic function and how these vessels differ from lung blood vasculature in their architecture, function, and response to injury in 3 domains: edema, inflammation, and thrombosis. We will review the rich historical anatomic literature that described the lung lymphatics in great detail and elucidated foundational discoveries that continue to shape our current understanding of the lung lymphatics, even in the molecular era. We conclude by highlighting key questions for the field of lung lymphatic biology.
Abohashem S, Saeed F, Münzel T
… +5 more, Al-Kindi SG, Foldyna B, Fayad ZA, Tawakol A, Osborne MT
Arterioscler Thromb Vasc Biol
· 2025 Dec · PMID 41064859
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Cardiovascular disease remains the leading cause of mortality worldwide, driven by risk factors that range from traditional (eg, hypertension, hyperlipidemia) to less recognized socioenvironmental contributors. These bro...Cardiovascular disease remains the leading cause of mortality worldwide, driven by risk factors that range from traditional (eg, hypertension, hyperlipidemia) to less recognized socioenvironmental contributors. These broader exposures include adverse socioeconomic status, air and noise pollution, attributes of the built environment, and ambient temperatures, among others, which exert complex mechanistic influences that often involve neural-autonomic-immune pathways that promote traditional cardiovascular disease risk factors and atherosclerosis. Advanced noninvasive imaging modalities, including positron emission tomography, computed tomography, magnetic resonance imaging, and ultrasound, allow for assessment of subclinical vascular changes, such as arterial inflammation and plaque burden, as well as assessments of changes in other organs, including the brain and inflammatory tissues, that associate with these exposures and have the potential to clarify the mechanisms of exposure-related pathology. This review synthesizes current evidence from multimodality imaging studies linking socioeconomic status, air pollution, noise, and other environmental exposures to imaging markers of cardiovascular disease. These findings suggest opportunities to deeply characterize underlying mechanisms, refine risk assessment, prioritize targeted interventions, and inform policies aimed at mitigating adverse exposures. Through this framework, we aim to catalyze a broader approach to preventing cardiovascular disease that recognizes the profound interplay among the social, environmental, and biological determinants of health.
Clark MD, Kim Y, Romero CA
… +12 more, Kang DW, Baek KI, Song EJ, Kellum CE, Bowman-Kirigin JA, Park C, Hung RC, Choi L, Kapoor V, Tsuji S, Pollock JS, Jo H
Arterioscler Thromb Vasc Biol
· 2025 Dec · PMID 41064858
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Aldosoky W, Abohashem S, Goudot G
… +10 more, Grewal SS, Qamar I, Hanlon E, Alani O, Bellinge J, Civieri G, Osborne MT, Dweck MR, Heidari P, Tawakol A
Arterioscler Thromb Vasc Biol
· 2025 Dec · PMID 41064857
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BACKGROUND: Inflammation potentiates aortic valve calcification (AVC). Ga-DOTATATE (gallium-68 DOTA-(Tyr³)-octreotate), a positron emission tomography tracer that binds to somatostatin receptors, provides a measure of ti...BACKGROUND: Inflammation potentiates aortic valve calcification (AVC). Ga-DOTATATE (gallium-68 DOTA-(Tyr³)-octreotate), a positron emission tomography tracer that binds to somatostatin receptors, provides a measure of tissue inflammation. However, the diagnostic and prognostic values of aortic valve (AV) Ga-DOTATATE uptake in AVC remain unexplored. Here, we tested whether AV Ga-DOTATATE uptake predicts the progression of AVC. METHODS: A total of 683 individuals (median age, 63 years; 46% male) underwent clinical Ga-DOTATATE positron emission tomography/computed tomography imaging from 2017 to 2023; 209 had follow-up imaging (median, 1.3 years interval). AV inflammation was measured as the maximum standardized uptake value of Ga-DOTATATE uptake within the AV on baseline positron emission tomography/computed tomography. AVC was quantified on baseline and follow-up computed tomography scans (Hounsfield units >130). AVC progression was assessed as the difference between baseline and follow-up AVC. Individuals with a square root difference of annualized AVC change ≥2.5 were characterized as progressors and <2.5 as nonprogressors. Demographic and clinical data were collected from medical records. RESULTS: Baseline AV Ga-DOTATATE uptake correlated with baseline AVC (standardized ρ=0.12; =0.002). Furthermore, baseline AV Ga-DOTATATE uptake associated with AVC progression (odds ratio [OR], 3.0 [95% CI, 1.4-6.4]; =0.004) and remained significant after separately adjusting for baseline AVC (OR, 3.1 [95% CI, 1.5-6.6]), sex (OR, 4.0 [95% CI, 1.7-9.0]), hypertension (OR, 2.8 [95% CI, 1.3-6.2]), diabetes (OR, 3.0 [95% CI, 1.4-6.4]), hyperlipidemia (OR, 2.4 [95% CI, 1.1-5.3]), smoking (OR, 3.1 [95% CI, 1.5-6.7]), chronic kidney disease (OR, 2.9 [95% CI, 1.4-6.3]), and body mass index (OR, 3.0 [95% CI, 1.4-6.3]), became insignificant when adjusting to age (OR, 1.9 [95% CI, 0.8-4.3]). CONCLUSIONS: Our study highlights the use of Ga-DOTATATE for assessing AV inflammation and predicting AVC progression. These findings underscore the role of inflammation in AVC progression and have potential implications for risk assessment and evaluating therapies in AV disease.
Jiang Z, Su L, Chen C
… +9 more, He R, Jiang L, Shu Y, Dai D, Wu M, Guo A, Liu J, Liu S, Liu Z
Arterioscler Thromb Vasc Biol
· 2025 Dec · PMID 41064856
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BACKGROUND: Ischemic diseases have become a major threat to global health, with endothelial cell (EC) damage closely associated with their pathogenesis and progression. Cell therapies targeting endothelial repair have th...BACKGROUND: Ischemic diseases have become a major threat to global health, with endothelial cell (EC) damage closely associated with their pathogenesis and progression. Cell therapies targeting endothelial repair have thus become a treatment approach of great interest, yet the procurement of clinically approved ECs for these applications has not been fully established. Modulating the expression of Atf3 (activating transcription factor 3) represents a potential strategy for deriving ECs from stem cells; however, its precise function in the development and differentiation of ECs from stem cells remains elusive. In the present study, we sought to elucidate the potential role of Atf3 in the differentiation of embryonic stem cells into ECs. METHODS: CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats/clustered regularly interspaced short palindromic repeat-associated 9) system was used to knockout Atf3 (Atf3KO [Atf3 knockout]) in mouse embryonic stem cells. EC differentiation was initially induced using the hanging drop method to promote embryoid bodies formation, followed by embryoid bodies attachment onto culture slides. The expression changes of EC markers during differentiation were assessed by RNA sequencing, Western blotting, immunofluorescence staining, flow cytometry, and reverse transcription quantitative polymerase chain reaction. Functional comparisons of differentiated ECs were performed by assessing LDL (low-density lipoprotein) uptake and NO production. Potential molecular mechanisms were further explored via bioinformatic analysis of RNA sequencing data. RESULTS: Atf3KO led to a significant upregulation in the expression levels of progenitor and mesoderm cell markers on days 3 and 6 of differentiation. By day 9, the expression of mature EC markers also exhibited a notable increase. Moreover, Atf3KO enhanced the functional properties of differentiated Atf3KO ECs. In addition, our findings revealed that the activation of the Rap1 (Ras-related protein 1) signaling pathway, triggered by Atf3KO, contributed to ECs development and maturation. CONCLUSIONS: Atf3KO directs embryonic stem cells toward the mesodermal lineage and activates the Rap1 signaling pathway, thereby promoting ECs development. These findings highlight a key role of Atf3 in regulating early stage of vascular endothelial development.
Krebs JR, Bellotti P, Ueland W
… +12 more, Valisno JAC, Joseph Manual Kollareth D, Sharma S, Su G, Hartman JB, Adithan A, Spinosa M, Kamat M, Garrett T, Cai G, Sharma AK, Upchurch GR
Arterioscler Thromb Vasc Biol
· 2025 Nov · PMID 41036562
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BACKGROUND: The pathogenesis of abdominal aortic aneurysm (AAA) formation involves vascular inflammation, thrombosis formation, and programmed cell death, leading to aortic remodeling. In this study, we deciphered the ro...BACKGROUND: The pathogenesis of abdominal aortic aneurysm (AAA) formation involves vascular inflammation, thrombosis formation, and programmed cell death, leading to aortic remodeling. In this study, we deciphered the role of ferroptosis, an excessive iron-mediated cell death in macrophages during aortic inflammation and vascular remodeling in AAA formation. METHODS: Single-cell RNA sequencing analysis was performed on the human AAA tissue database. AAAs were induced in male and female C57BL/6 (wild-type) mice using 2 models with topical elastase or elastase+β-aminopropionitrile, with or without liproxstatin-1, a specific ferroptosis inhibitor, treatment. Aortic diameter, cytokine expression, histology, hallmarks of ferroptosis such as lipid peroxidation and glutathione, and lipid analysis using mass spectrometry were measured in aortic tissue extracts. In vitro studies deciphered the crosstalk of macrophages and smooth muscle cells and analyzed ferroptosis and MMP (matrix metalloproteinase) expressions. RESULTS: Single-cell RNA sequencing analysis demonstrated significant differences in ferroptosis-related genes in macrophages from human AAAs compared with control aortic tissue. Using 2 established murine models of AAA and aortic rupture in wild-type mice, we observed that treatment with liproxstatin-1 significantly attenuated aortic diameter, proinflammatory cytokine production, immune cell infiltration (neutrophils and macrophages), elastic fiber disruption, and increased smooth muscle cell α-actin expression compared with untreated mice. Lipidomic analysis using mass spectrometry shows a significant increase in ceramides and a decrease in intact lipid species levels in murine AAA tissue compared with controls in the murine AAA model. Mechanistically, in vitro studies demonstrate that liproxstatin-1 treatment of macrophages mitigated ferroptosis and MMP9 expression, as well as the crosstalk with aortic smooth muscle cells by downregulating MMP2 secretion. CONCLUSIONS: Taken together, this study demonstrates that pharmacological inhibition by liproxstatin-1 mitigates macrophage-dependent ferroptosis, contributing to the inhibition of aortic inflammation and remodeling during AAA formation.