The effects of calcitonin gene-related peptide (CGRP) on atherosclerosis remain unclear. We used apolipoprotein E-deficient (ApoE-/-) mice to generate double-knockout ApoE-/-:CGRP-/- mice lacking alpha CGRP. ApoE-/-:CGRP-/- mice exhibited larger atherosclerotic plaque areas, peritoneal macrophages with enhanced migration functions, and elevated levels of the inflammatory cytokine tumor necrosis factor (TNF)-⍺. Thus, we also explored whether inhibiting TNF-⍺ could improve atherosclerosis in ApoE-/-:CGRP-/- mice by administering etanercept intraperitoneally once a week (5 mg/kg) alongside a high-fat diet for 2 weeks. This treatment led to significant reductions in aortic root lesion size, atherosclerotic plaque area and macrophage migration in ApoE-/-:CGRP-/- mice compared with mice treated with human IgG (5 mg/kg). We further examined whether results observed in ApoE-/-:CGRP-/- mice could similarly be obtained by administering a humanized monoclonal CGRP antibody, galcanezumab, to ApoE-/- mice. ApoE-/- mice were subcutaneously administered galcanezumab at an initial dose of 50 mg/kg, followed by a dose of 30 mg/kg in the second week. Galcanezumab administration did not affect systolic blood pressure, serum lipid levels, or macrophage migration but led to a significant increase in lipid deposition at the aortic root. These findings suggest that alpha CGRP plays a critical role in inhibiting the progression of atherosclerosis.

The onset and progression of atherosclerosis are closely linked to the involvement of macrophages. While the contribution of NLRP3 inflammasome activation to the creation of a local highly inflammatory microenvironment is well recognized, the precise triggers remain unclear. In this study, we aimed to investigate the regulatory mechanism of NLRP3 inflammasome activation in response to hypoxia-induced glycolysis involving PFKFB3 in the development of atherosclerosis. To develop an atherosclerosis model, we selected ApoE knockout mice treated with a high-fat western diet. We then quantified the expression of HIF-1α, PFKFB3, and NLRP3. In addition, we administered the PFKFB3 inhibitor PFK158 during atherosclerosis modeling. The glycolytic activity was subsequently determined through 18F-FDG micro-PET/CT, ex vivo glucose uptake, and ECAR analysis. Furthermore, we employed lipopolysaccharide (LPS) and TNF-α to induce the differentiation of bone marrow-derived macrophages (BMDMs) into M1-like phenotypes under both hypoxic and normoxic conditions. Our histological analyses revealed the accumulation of PFKFB3 in human atherosclerotic plaques, demonstrating colocalization with NLRP3 expression and macrophages. Treatment with PFK158 reduced glycolytic activity and NLRP3 inflammasome activation, thereby mitigating the occurrence of atherosclerosis. Mechanistically, hypoxia promoted glycolytic reprogramming and NLRP3 inflammasome activation in BMDMs. Subsequent blocking of either HIF-1α or PFKFB3 downregulated the NLRP3/Caspase-1/IL-1β pathway in hypoxic BMDMs. Our study demonstrated that the HIF-1α/PFKFB3/NLRP3 axis serves as a crucial mechanism for macrophage inflammation activation in the emergence of atherosclerosis. The therapeutic potential of PFKFB3 inhibition may represent a promising strategy for atheroprotection.

BACKGROUND: Atherosclerosis (AS) is the most prevalent cardiovascular disease, with an exceptionally high burden. High-fat diet (HFD) is a popular diet behavior, whereas low-dose radiation (LDR) is an environmental physical factor. There is evidence to suggest that an HFD may exacerbate the onset of atherosclerosis. Whether the combination effect of HFD and LDR would have potential on atherosclerosis development remains incompletely unclear.
METHODS: In this study, ApoE-/- mice were used as atherosclerosis model animals to investigate the combination effects of HFD and LDR (10 × 0.01Gy, or 20 × 0.01Gy) on vascular lesions. Doppler ultrasound imaging, H&E staining, oil red O staining, western blotting, and immunohistochemistry (IHC) were used to assess the pro-atherosclerotic effects. LC-MS was used to detect the non-targeted lipidomic.
RESULTS: Long-term exposure of low-dose radiation at an accumulated dose of 0.2Gy significantly increased the occurrence of vascular stiffness and the aortic lesion in ApoE-/- mice. The synergistic effect of HFD and LDR was observed in the development of atherosclerosis, which might be linked to both the dysbiosis of lipid metabolism and the stimulation of the inflammatory signaling system. Moreover, LDR but not HFD can activate the cGAS-STING signaling through increasing the yield of cytosolic mitochondrial DNAs as well as the expression of cGAS protein. The activation of cGAS-STING signal triggers the release of IFN-α/-β, which functions as an inflammatory amplifier in the formation of atherosclerotic plaque.
CONCLUSIONS: The current study offers fresh insights into the risks and mechanism that underlie the development of atherosclerosis by LDR, and there is a combination effect of LDR and HFD with the involvement of cGAS-STING signal pathway.

BACKGROUND: Qinghao-Biejia herb pair (QB) is the core herb pair of \"Jieduquyuziyin prescription\" and is one of the commonly used herb pairs for the clinical treatment of systemic lupus erythematosus (SLE). Previous studies have shown that QB reduces the expression of inflammatory cytokines like IL-6 and TNF-α in the serum and kidney of MRL/lpr mice. Additionally, it inhibits the expression of TLR4 and MyD88 in the kidney and aorta and reduces the deposition of renal complement C3 and aortic plaque after treatment. These findings suggest that QB has a preventive and therapeutic effect on lupus rats.
OBJECTIVE: This study sought to investigate the mechanisms underlying the anti-SLE combined with atherosclerosis activity of the Qinghao-Biejia herb pair.
METHODS: Drug targets for QB were identified using the HERB database, while targets associated with SLE and atherosclerosis were retrieved from the GeneCards database. The intersection of these drug and disease targets was then analyzed using a protein-protein interaction (PPI) network with GO and KEGG pathway enrichment analysis. In vivo, apolipoprotein E-deficient (ApoE-/-) mice were induced to develop SLE-AS by intraperitoneal injection of pristane and continued feeding of a high-fat diet. The changes in relevant indexes were observed after 12 weeks of gavage treatment with hydroxychloroquine, QB, Q (Qinghao alone), and B (Biejia alone). Bone marrow-derived macrophages from ApoE-/- mice and Raw 264.7 macrophages were used to explore the mechanisms of QB treatment.
RESULTS: The levels of inflammatory cytokines in serum and pathological liver changes in mice were improved to varying degrees in the treatment groups. Additionally, there was a reduction in aortic atheromatous plaque formation and some improvement in cholesterol efflux. Furthermore, QB suppressed the expression of inflammatory cytokines in M1 macrophages, suggesting a role in regulating macrophage polarization.
CONCLUSIONS: QB demonstrates clear efficacy for treating SLE-AS, and its therapeutic mechanism may involve the regulation of macrophage phenotypes by promoting cholesterol efflux.

BACKGROUND: Hyperlipidemia damages vascular wall and serves as a foundation for diseases such as atherosclerosis, hypertension and stiffness. The NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome is implicated in vascular dysfunction associated with hyperlipidemia-induced vascular injury. Sodium tanshinone IIA sulfonate (STS), a well-established cardiovascular protective drug with recognized anti-inflammatory, antioxidant, and vasodilatory properties, is yet to be thoroughly investigated for its impact on vascular relaxant imbalance induced by hyperlipidemia.
METHODS: In this study, we treated ApoE-knockout (ApoE-/-) mouse with STS and assessed the activation of the NLRP3 inflammasome, expression of MMP2/9, integrity of elastic fibers, and vascular constriction and relaxation.
RESULTS: Our findings reveal that STS intervention effectively preserves elastic fibers, significantly restores aortic relaxation function in ApoE-/- mice, and reduces their excessive constriction. Furthermore, STS inhibits the phosphorylation of spleen tyrosine kinase (SYK), suppresses NLRP3 inflammasome activation, and reduces MMP2/9 expression.
CONCLUSIONS: These results demonstrate that STS protects vascular relaxation against hyperlipidemia-induced damage through modulation of the SYK-NLRP3 inflammasome-MMP2/9 pathway. This research provides novel insights into the mechanisms underlying vascular relaxation impairment in a hyperlipidemic environment and uncovers a unique mechanism by which STS preserves vascular relaxation, offering valuable foundational research evidence for its clinical application in promoting vascular health.

Atherosclerosis (AS) is the main pathological basis of cardiovascular diseases such as coronary heart disease. Black phosphorus quantum dots (BPQDs) are a novel nanomaterial with good optical properties and biocompatibility, which was applied in the treatment of AS in mice, with good results shown in our previous study. In this study, BPQDs were injected into high-fat diet-fed apolipoprotein E knockout mice as a preventive drug for 12 weeks. Simvastatin, a classic preventive drug for AS, was used as a control to verify the preventive effect of BPQDs. The results showed that after preventive treatment with BPQDs, the plaque area in mice was significantly reduced, the vascular elasticity was increased, and serum lipid levels were significantly lower than those in the model group. To explore the mechanism, macrophages were induced to become foam cells using oxidized low-density lipoprotein. We found that BPQDs treatment could increase cell autophagy, thereby regulating intracellular lipid metabolism. Taken together, these data revealed that BPQDs may serve as a functional drug in preventing the development of AS.

Vulnerable atherosclerotic plaque rupture, the leading cause of fatal atherothrombotic events, is associated with an increased risk of mortality worldwide. Peroxisome proliferator-activated receptor delta (PPARδ) has been shown to modulate vascular smooth muscle cell (SMC) phenotypic switching, and, hence, atherosclerotic plaque stability. Melatonin reportedly plays a beneficial role in cardiovascular diseases; however, the mechanisms underlying improvements in atherosclerotic plaque vulnerability remain unknown. In this study, we assessed the role of melatonin in regulating SMC phenotypic switching and its consequential contribution to the amelioration of atherosclerotic plaque vulnerability and explored the mechanisms underlying this process. We analyzed features of atherosclerotic plaque vulnerability and markers of SMC phenotypic transition in high-cholesterol diet (HCD)-fed apolipoprotein E knockout (ApoE-/-) mice and human aortic SMCs (HASMCs). Melatonin reduced atherosclerotic plaque size and necrotic core area while enhancing collagen content, fibrous cap thickness, and smooth muscle alpha-actin positive cell coverage on the plaque cap, which are all known phenotypic characteristics of vulnerable plaques. In atherosclerotic lesions, melatonin significantly decreased the synthetic SMC phenotype and KLF4 expression and increased the expression of PPARδ, but not PPARα and PPARγ, in HCD-fed ApoE-/- mice. These results were subsequently confirmed in the melatonin-treated HASMCs. Further analysis using PPARδ silencing and immunoprecipitation assays revealed that PPARδ plays a role in the melatonin-induced SMC phenotype switching from synthetic to contractile. Collectively, we provided the first evidence that melatonin mediates its protective effect against plaque destabilization by enhancing PPARδ-mediated SMC phenotypic switching, thereby indicating the potential of melatonin in treating atherosclerosis.

BACKGROUND: Atherosclerosis is highly prevalent in people with chronic kidney disease (CKD), including those receiving peritoneal dialysis (PD). Although it is lifesaving, PD induces profound systemic inflammation, which may aggravate atherosclerosis. Therefore, the hypothesis is that this PD-induced inflammation aggravates atherosclerosis via immune cell activation.
RESULTS: ApoE-/- mice were subjected to a 5/6 nephrectomy to induce CKD. Three weeks later, mice were fed a high-cholesterol diet. Half of the nephrectomized mice then received daily peritoneal infusions of 3.86% Physioneal for 67 further days (CKD+PD) until the end of the experiment, and were compared with mice without CKD. Sham operated and PD-only mice were additional controls. CKD+PD mice displayed more severe atherosclerotic disease than control mice. Plaque area increased, and plaques were more advanced with a vulnerable phenotype typified by decreased collagen content and decreased fibrous cap thickness. Increased CD3+ T-cell numbers were present in plaques and perivascular adipose tissue of CKD and CKD+PD mice. Plaques of CKD+PD mice contained more iNOS+ immune cells. Spleens of CKD+PD mice showed more CD4+ central memory, terminally differentiated type 1 T-helper (Th1), Th17, and CX3C motif chemokine receptor 1+ (CX3CR1) CD4+ T-cells with less regulatory and effector T-cells.
CONCLUSIONS: PD-fluid exposure in uremic mice potentiates systemic and vascular T-cell-driven inflammation and aggravates atherosclerosis. PD polarized CD4+ T-cells toward an inflammatory Th1/Th17 phenotype, and increased CX3CR1+ CD4+ T-cells, which are associated with vascular homing in CKD-associated atherosclerosis. Targeting CD4+ T-cell activation and CX3CR1+ polarization has the potential to attenuate atherosclerosis in PD patients.

UNASSIGNED: The long isoform of the Wnk1 (with-no-lysine [K] kinase 1) is a ubiquitous serine/threonine kinase, but its role in vascular smooth muscle cells (VSMCs) pathophysiology remains unknown.
UNASSIGNED: AngII (angiotensin II) was infused in Apoe-/- to induce experimental aortic aneurysm. Mice carrying an Sm22-Cre allele were cross-bred with mice carrying a floxed Wnk1 allele to specifically investigate the functional role of Wnk1 in VSMCs.
UNASSIGNED: Single-cell RNA-sequencing of the aneurysmal abdominal aorta from AngII-infused Apoe-/- mice revealed that VSMCs that did not express Wnk1 showed lower expression of contractile phenotype markers and increased inflammatory activity. Interestingly, WNK1 gene expression in VSMCs was decreased in human abdominal aortic aneurysm. Wnk1-deficient VSMCs lost their contractile function and exhibited a proinflammatory phenotype, characterized by the production of matrix metalloproteases, as well as cytokines and chemokines, which contributed to local accumulation of inflammatory macrophages, Ly6Chi monocytes, and γδ T cells. Sm22Cre+Wnk1lox/lox mice spontaneously developed aortitis in the infrarenal abdominal aorta, which extended to the thoracic area over time without any negative effect on long-term survival. AngII infusion in Sm22Cre+Wnk1lox/lox mice aggravated the aortic disease, with the formation of lethal abdominal aortic aneurysms. Pharmacological blockade of γδ T-cell recruitment using neutralizing anti-CXCL9 (anti-CXC motif chemokine ligand 9) antibody treatment, or of monocyte/macrophage using Ki20227, a selective inhibitor of CSF1 receptor, attenuated aortitis. Wnk1 deletion in VSMCs led to aortic wall remodeling with destruction of elastin layers, increased collagen content, and enhanced local TGF-β (transforming growth factor-beta) 1 expression. Finally, in vivo TGF-β blockade using neutralizing anti-TGF-β antibody promoted saccular aneurysm formation and aorta rupture in Sm22 Cre+ Wnk1lox/lox mice but not in control animals.
UNASSIGNED: Wnk1 is a key regulator of VSMC function. Wnk1 deletion promotes VSMC phenotype switch toward a pathogenic proinflammatory phenotype, orchestrating deleterious vascular remodeling and spontaneous severe aortitis in mice.

UNASSIGNED: Aging increases the risk of atherosclerotic vascular disease and its complications. Macrophages are pivotal in the pathogenesis of vascular aging, driving inflammation and atherosclerosis progression. NOX4 (NADPH oxidase 4) expression increases with age, correlating with mitochondrial dysfunction, inflammation, and atherosclerosis. We hypothesized that the NOX4-dependent mitochondrial oxidative stress promotes aging-associated atherosclerosis progression by causing metabolic dysfunction and inflammatory phenotype switch in macrophages.
UNASSIGNED: We studied atherosclerotic lesion morphology and macrophage phenotype in young (5-month-old) and aged (16-month-old) Nox4 -/-/Apoe -/- and Apoe -/- mice fed Western diet.
UNASSIGNED: Young Nox4-/-/Apoe-/- and Apoe-/- mice had comparable aortic and brachiocephalic artery atherosclerotic lesion cross-sectional areas. Aged mice showed significantly increased lesion area compared with young mice. Aged Nox4-/-/Apoe-/- had significantly lower lesion areas than Apoe-/- mice. Compared with Apoe-/- mice, atherosclerotic lesions in aged Nox4-/-/Apoe-/- showed reduced cellular and mitochondrial ROS and oxidative DNA damage, lower necrotic core area, higher collagen content, and decreased inflammatory cytokine expression. Immunofluorescence and flow cytometry analysis revealed that aged Apoe-/- mice had a higher percentage of classically activated pro-inflammatory macrophages (CD38+CD80+) in the lesions. Aged Nox4-/-/Apoe-/- mice had a significantly higher proportion of alternatively activated pro-resolving macrophages (EGR2+/CD163+CD206+) in the lesions, with an increased CD38+/EGR2+ cell ratio compared with Apoe-/- mice. Mitochondrial respiration assessment revealed impaired oxidative phosphorylation and increased glycolytic ATP production in macrophages from aged Apoe-/- mice. In contrast, macrophages from Nox4-/-/Apoe-/- mice were less glycolytic and more aerobic, with preserved basal and maximal respiration and mitochondrial ATP production. Macrophages from Nox4-/-/Apoe-/- mice also had lower mitochondrial ROS levels and reduced IL1β secretion; flow cytometry analysis showed fewer CD38+ cells after IFNγ+LPS treatment and more EGR2+ cells after IL4 treatment than in Apoe-/- macrophages. In aged Apoe-/- mice, inhibition of NOX4 activity using GKT137831 significantly reduced macrophage mitochondrial ROS and improved mitochondrial function, resulting in decreased CD68+CD80+ and increased CD163+CD206+ lesion macrophage proportion and attenuated atherosclerosis.
UNASSIGNED: Our findings suggest that increased NOX4 in aging drives macrophage mitochondrial dysfunction, glycolytic metabolic switch, and pro-inflammatory phenotype, advancing atherosclerosis. Inhibiting NOX4 or mitochondrial dysfunction could alleviate vascular inflammation and atherosclerosis, preserving plaque integrity.