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BACKGROUND: Ferroptosis is an iron-dependent regulatory form of cell death characterized by iron-dependent lipid peroxidation. Long-chain acyl-coenzyme A synthase 4 (ACSL4) is involved in the formation of lipid peroxidation substrates, thereby resulting in ferroptosis. Recent studies have shown that ACSL4- mediated ferroptosis plays a key role in atherosclerotic cardiovascular disease. OBJECTIVE: To summarize the structural function and regulatory mechanism of ACSL4 and its potential molecular mechanism mediating ferroptosis, and to elaborate the application of ACSL4 driving ferroptosis in atherosclerosis, ischemic stroke and myocardial infarction, in order to provide a new therapeutic strategy for the treatment of atherosclerotic cardiovascular diseases. METHODS: Relevant literature was searched in PubMed database from database inception to August 2023 using the keywords of “atherosclerosis, ferroptosis, long-chain acyl-coenzyme A synthase 4, ACSL4, glutathione peroxidase 4, ischemic stroke, myocardial infarction, endothelial cell, smooth muscle cells, foam cell.” Finally, 76 documents were included for review and analysis. RESULTS AND CONCLUSION: ACSL4 participates in the formation of coenzyme derivatives of polyunsaturated fatty acids and inserts them into phospholipids to provide substrates for lipid peroxidation, the core mechanism of iron death. Among the regulatory factors of ACSL4 expression, integrin α6β4, intracellular vesicular transport factor p115, and zinc lipoprotein A20 negatively regulate its expression. Meanwhile, multiple miRs down-regulate its expression by binding to 3’-UTR. On the contrary, up-regulation of ACSL4 is mostly regulated by transcription factors. ACSL4-dependent production of phospholipids containing polyunsaturated fatty acids is an essential prerequisite for lipid peroxidation and ferroptosis. Moreover, ACSL4 and glutathione peroxidase 4 are mutually dependent as positive and negative regulators of ferroptosis, and their specific mechanisms remain to be further studied. ACSL4-mediated ferroptosis is involved in the pathological mechanism of atherosclerosis, ischemic stroke, and myocardial infarction. Endothelial cell injury in atherosclerosis is closely related to ACSL4-mediated ferroptosis, but there are no reports on the involvement of ACSL4 in foam cell formation, smooth muscle cell phenotype transformation, and calcification. ACSL4 has become a research hotspot as a biomarker and potential target of ferroptosis. Targeting ACSL4 to inhibit ferroptosis may become a new direction for the treatment of atherosclerotic cardiovascular diseases. However, there are few studies on drugs inhibiting ACSL4, and further studies are needed in the future. [ABSTRACT FROM AUTHOR]

Background: Macrophage-derived foam cells are essential in the progression of atherosclerosis (AS). Based on our previous study, the Huxin Formula (HXF), a traditional Chinese medicine formula, demonstrates potential in anti-atherosclerosis. Nevertheless, it is still unknown how HXF affects the formation of foam cells derived from THP-1 macrophages. Purpose: This research aims to examine the preventive role of HXF in the development of foam cells and its underlying molecular mechanism. Methods: THP-1 derived macrophages and THP-1 cells overexpressing LOX-1 (LV-OLR1) were exposed to ox-LDL to establish foam cell models, and then treated with HXF. Meantime, Oil red O staining was used to detect lipid droplet production. ELISA kit was performed to measure intracellular levels of IL-6 and TGF-β. RT-qPCR and Western Blot were then utilized to determine the LOX-1 and NF-κB mRNA/protein levels. Results: The findings indicated that HXF treatment potently reduced the lipid accumulation, downregulated IL-6 levels and upregulated TGF-β levels. However, this impact was almost reversed when LOX-1 was overexpressed in THP-1 cells stimulated with ox-LDL. Moreover, THP-1 were treated with HXF markedly reduced the levels of LOX-1 and NF-κB mRNA/protein, whereas overexpressing LOX-1 significantly reversed this effect. Conclusion: HXF reduced the formation of foam cell in ox-LDL-stimulated THP-1 macrophages via inhibiting lipid accumulation and inflammation through regulating the LOX-1/ NF-κB pathway. These present findings further indicate a potential beneficial role of HXF in ameliorating atherosclerosis and foam cell formation, while provide a novel potential therapeutic strategy for preventing atherosclerosis. [ABSTRACT FROM AUTHOR]

Macrophage-derived lipid-laden foam cells from the subendothelium play a crucial role in the initiation and progression of atherosclerosis. However, the molecule mechanism that regulates the formation of foam cells is not completely understood. Here, we found that SLAMF7 was upregulated in mice bone marrow–derived macrophages and RAW264.7 cells stimulated with oxidized low-density lipoprotein (ox-LDL). SLAMF7 promoted ox-LDL-mediated macrophage lipid accumulation and M1-type polarization. SLAMF7 deficiency reduced serum lipid levels and improved the lesions area of carotid plaque and aortic arch in high-fat diet-fed ApoE−/− mice. In response to ox-LDL, SLAMF7 downregulated NR4A1 and upregulated RUNX3 through transcriptome sequencing analysis. Overexpression NR4A1 reversed SLAMF7-induced lipid uptake and M1 polarization via inhibiting RUNX3 expression. Furthermore, RUNX3 enhanced foam cell formation and M1-type polarization. Taken together, the study suggested that SLAMF7 play contributing roles in the pro-atherogenic effects by regulating NR4A1-RUNX3. [ABSTRACT FROM AUTHOR]

In our search for bioactive components, various chromatographic separations of the organic fractions from Filipendula glaberrima leaves led to the isolation of a new ellagitannin and a triterpenoid, along with 26 known compounds. The structures of the isolates were determined based on their spectroscopic properties and chemical evidence, which were then evaluated for their antioxidant activities, inhibitory activities on 3-hydroxy-3-methylglutaryl-coenzyme A reductase, and foam cell formation in THP-1 cells to prevent atherosclerosis. Rugosin B methyl ester (1) showed the best HMG-CoA reductase inhibition and significantly reduced ox-low-density lipoprotein-induced THP-1 macrophage-derived foam cell formation at 25 µM. In addition, no cytotoxicity was observed in THP-1 cells at 50 μg/mL of all extracts in the macrophage foam cell formation assay. Therefore, F. glaberrima extract containing 1 is promising in the development of dietary supplements due to its potential behavior as a novel source of nutrients for preventing and treating atherosclerosis. [ABSTRACT FROM AUTHOR]

Objective To explore the inhibitory effect of Danyu formula on foam cell formation in rat aortic endothelial cells (RAECs) induced by oxidized low-density lipoprotein (Ox-LDL) and its possible mechanism. Methods RAECs were randomly divided into the control group, model group, low-dose Danyu formula group, medium-dose Danyu formula group, and high-dose Danyu formula group, and atorvastatin group. Except for the control group, RAECs in the rest of the five groups were treated with 200 µg/mL Ox-LDL for 72 h, and the low-, medium-, and high-dose Danyu formula groups and the atorvastatin group were treated with 50, 100, and 200 µg/L Danyu formula solution and 300 µg/L atorvastatin, respectively. After 24, 48, and 72 h of treatment, the cell proliferation ability was detected by CCK-8. The foam formation was observed by oil red O staining in the cells treated for 72 h in each group. The protein levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), malondialdehyde (MDA), and superoxide dismutase (SOD) in the cell supernatant were measured by ELISA. The reactive oxygen species (ROS) content was determined by the 2-hydroxyethylfluorescein oxidation fluorescence reaction. The relative mRNA expression levels of protein tyrosine kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) were detected by real-time fluorescence quantitative PCR. The relative protein expression levels of JAK2, p-JAK2, STAT3, and p-STAT3 were determined by Western blotting, and the ratios of p-JAK2/JAK2 and p-STAT3/STAT3 were calculated. Results Compared with the control group, the OD values of the model group decreased at the same time points (all P<0. 05). Compared with the model group at the same time points, the OD values of the high-dose Danyu formula group and the atorvastatin group increased, and the OD values of the mediumdose Danyu formula group increased at 48 and 72h (all P<0. 05). The control group cells did not exhibit foam formation, while the cells in the model group showed obvious foam formation. Compared with the model group, the foam formation in the atorvastatin group and the high-, medium-, and low-dose Danyu formula groups improved, with the high-dose Danyu formula group and the atorvastatin group showing the most significant improvement, almost approaching the control group. Compared with the control group, the levels of TNF-α, IL-6, ROS, JAK2 mRNA expression, STAT3 mRNA expression, p-JAK2/JAK2, p-STAT3/STAT3, and MDA in the cell supernatant of the model group increased, while the expression of SOD in the cell supernatant decreased (all P<0. 05). Compared with the model group, the levels of TNF-α, IL-6, ROS, JAK2 mRNA expression, STAT3 mRNA expression, p-JAK2/JAK2, p-STAT3/STAT3, and MDA in the cell supernatant of the atorvastatin group and the high-, medium-, and low-dose Danyu formula groups decreased, while the expression of SOD in the cell supernatant increased, with the most significant changes in the atorvastatin group and the high-dose Danyu formula group (all P<0. 05). There were no significant differences in the relative protein expression levels of JAK2 or STAT3 among these groups (all P>0. 05). Conclusion The high-dose, medium-dose, and low-dose Danyu formula can inhibit foam formation in Ox-LDL-induced RAECs, with the significant effect in the high-dose Danyu formula, and its mechanism may be related to the inhibition of inflammatory reaction, oxidative stress response, and JAK2/STAT3 signaling pathway. [ABSTRACT FROM AUTHOR]

Abstract Aims Atherosclerosis is a huge threaten to the human health, C1q/TNF‐related protein 9 (CTRP9) has been previously reported possessing vascular protective functions. Our study is aimed to reveal the mechanism of the regulative effects of CTRP9 on the foam cell formation. Methods and results Primary human macrophages were isolated from human monocytes donated by healthy volunteers. CCK‐8 assay was performed for determining the cell viability. Oil Red O staining was employed for measuring the lipid accumulation. Cholesterol ester and cholesterol concentration were detected by commercial kits for evaluating the intracellular cholesterol. Ubiquitination assay was performed to reveal the ubiquitination level of CD36, cycloheximide assay was applied for determining the half‐life of CD36 protein. Quantitative real‐time PCR and western blot assays were performed for detecting the mRNA and protein expression. Pre‐treatment with CTRP9 in primary human macrophages markedly suppressed the cholesterol accumulation concentration after oxidized low‐density lipoprotein treatment. CD36 was significantly increased after oxidized low‐density lipoprotein exposure while was reduced by CTRP9 treatment. Up‐regulation of CD36 significantly reversed the CTRP9‐mediated protective effects in foam cells. The differential expression levels of several deubiquitinating enzymes preliminarily indicated that USP11 was obviously decreased after CTRP9 treatment. USP11 knockdown decreased the CD36 protein expression and pre‐treatment with 10 μg/mL MG132 significantly maintained the CD36 level from USP11 knock down. Up‐regulation of CD36 reversed the alterations on the cholesterol metabolism caused by CTRP9 or USP11 knockdown. Conclusions CTRP9 regulates the USP11/CD36 axis to protect the macrophages form transforming into foam cells by suppressing intracellular lipid and cholesterol accumulation, which is a potential therapeutic agent for atherosclerosis.

In atherosclerosis, macrophage-derived foam cell formation is considered to be a hallmark of the pathological process; this occurs via the uptake of modified lipoproteins. In the present study, we aim to determine the role of transaldolase in foam cell formation and atherogenesis and reveal the mechanisms underlying its role. Bone marrow-derived macrophages (BMDMs) isolated from mice successfully form foam cells after treatment with oxidized low-density lipoprotein (80 μg/mL). Elevated transaldolase levels in the foam cell model are assessed by quantitative polymerase chain reaction and western blot analysis. Transaldolase overexpression and knockdown in BMDMs are achieved via plasmid transfection and small interfering RNA technology, respectively. We find that transaldolase overexpression effectively attenuates, whereas transaldolase knockdown accelerates, macrophage-derived foam cell formation through the inhibition or activation of cholesterol uptake mediated by the scavenger receptor cluster of differentiation 36 (CD36) in a p38 mitogen-activated protein kinase (MAPK) signaling-dependent manner. Transaldolase-mediated glutathione (GSH) homeostasis is identified as the upstream regulator of p38 MAPK-mediated CD36-dependent cholesterol uptake in BMDMs. Transaldolase upregulates GSH production, thereby suppressing p38 activity and reducing the CD36 level, ultimately preventing foam cell formation and atherosclerosis. Thus, our findings indicate that the transaldolase-GSH-p38-CD36 axis may represent a promising therapeutic target for atherosclerosis.

Seafood consumption has a great demand in the world due to its array of health benefits. Marine-derived bioactive peptides and hydrolysates have been found to have multifunctional health benefits. In this study, inhibitory effects of blue mussel hydrolysates, including blue mussel Alcalase® hydrolysate (BMAH) and blue mussel-pepsin hydrolysate (BMPH), on foam cell formation in ox-LDL-induced RAW264.7 macrophages were investigated. BMAH inhibited lipid accumulation evidenced by Oil Red O staining. Its inhibitory effect was higher than that of BMPH. BMAH positively modulated levels of total cholesterol, free cholesterol, and cholesterol ester in model cells. It suppressed cholesterol influx by downregulating SR-A1 and CD36 expression, but increased cholesterol efflux by upregulating ABCA-1 and ABCG-1 expression. In addition, ox-LDL treated RAW264.7 macrophages produced inflammatory responses and oxidative stress during foam cell formation. However, BMAH treatment significantly ameliorated proinflammatory cytokines production (TNF-α, IL-6, and IL-1β) by inhibiting NF-κB nuclear translocation. Moreover, BMAH inhibited intracellular reactive oxygen species (ROS) generation by activating HO-1/Nrf2 signaling pathway in ox-LDL-treated RAW264.7 macrophages. Taken together, these results suggest that BMAH might be useful as an ingredient of functional foods to ameliorate the development of atherosclerosis. [Display omitted] • Anti-atherosclerotic bioactive peptides were prepared from blue mussel protein. • Blue mussel Alcalase® hydrolysate (BMAH) inhibited oxidized LDL-induced foam cell formation. • Inhibition of BMAH against foam cell formation are associated with cholesterol metabolism. • BMAH attenuated inflammation and oxidative stress in oxidized LDL-induced foam cell formation. • BMAH inhibited NF-κB signaling and activated HO-1/Nrf2 signaling. [ABSTRACT FROM AUTHOR]

Cutaneous xanthoma consist of foam cells that originate from monocytes or macrophages and accumulate in perivascular areas of the skin. The main component of these cells is oxidized low‐density lipoprotein (oxLDL). In this study, we show that mast cells surround the accumulated foam cells, suggesting their involvement in xanthoma formation. Coculture of THP‐1 or U937 monocytes with the human mast cell line LUVA upregulated their uptake of oxLDL. Positive staining for intracellular cell adhesion molecule‐1 (ICAM‐1) at the borders between mast cells and foam cells was seen in pathological specimens of the most common cutaneous xanthoma, xanthelasma palpebrarum, and in cocultures. In the latter, ICAM1 messenger RNA levels were upregulated. The administration of anti‐ICAM‐1 blocking antibody inhibited the increase in oxLDL uptake by THP‐1 or U937 monocytes cocultured with LUVA. Taken together, these results suggest a role for mast cells in the formation of xanthelasma palpebrarum and the involvement of ICAM‐1 in this process. [ABSTRACT FROM AUTHOR]

Abstract Background Gastrodin is an effective polyphenol extracted from Chinese natural herbal Gastrodiae elata Blume, which exhibits antioxidant and anti-inflammatory effects. It has been reported to benefit neurodegenerative diseases, but the effect of Gastrodin on atherosclerosis and the underlying mechanisms remain elusive. The aim of this study is to investigate the function and mechanism of Gastrodin in atherosclerosis. Methods Atherosclerosis mouse model was established by fed low density lipoprotein receptor-deficient (Ldlr −/− ) mice with a high fat diet (HFD, 20% fat and 0.5 cholesterol) for 8 weeks and Gastrodin was administered daily via oral gavage. Plasma lipid levels were measured using commercial kits. En face and aortic sinus lipid accumulation were analyzed with Oil Red O staining. In vitro cell models using foam cell formation model and classical atherosclerosis inflammation model, macrophages were incubated with oxygenized low-density lipoproteins (ox-LDL) or lipopolysaccharide (LPS) in the presence of different concentration of Gastrodin or vehicle solution. Foam cell formation and cellular lipid content were evaluated by Oil Red O staining and intracellular lipids extraction analysis. Gene expression and proteins related to cholesterol influx and efflux were examined by quantitative reverse transcription PCR (RT-qPCR) and western blotting analysis. Furthermore, the effect of Gastrodin on LPS induced macrophage inflammatory responses and NF-κB pathway were evaluated by RT-qPCR and western blotting analysis. Results Gastrodin administration reduced the body weight, plasma lipid levels in Ldlr −/− mice after fed a high fat diet. Oil Red O staining showed Gastrodin-treated mice displayed less atherosclerosis lesion area. Furthermore, Gastrodin treatment significantly ameliorated ox-LDL-induced macrophage-derived foam cells formation through suppressing genes expression related to cholesterol efflux including scavenger receptor class B and ATP-binding cassette transporter A1. Moreover, Gastrodin markedly suppressed pro-inflammatory cytokines secretion and LPS induced inflammatory response in macrophage through downregulating NF-κB pathway. Conclusions Our study demonstrated that Gastrodin attenuates atherosclerosis by suppressing foam cells formation and LPS-induced inflammatory response and represents a novel therapeutic target for the treatment of atherosclerosis.

Abstract Background Atherosclerosis is driven by synergistic interactions between pathological biomechanical and lipid metabolic factors. Long noncoding RNAs (LncRNAs) have been implicated in atherogenesis. The purpose of this study was to investigate the potential mechanism of lncRNA AI662270 on macrophage cholesterol transport in atherosclerosis. Methods Apolipoprotein E deficiency (ApoE −/−) mice were fed a high fat diet for 16 weeks to construct atherosclerotic model, and the mice were injected with recombinant lentivirus carrying AI662270 gene to overexpress AI662270. Macrophages were cleared by liposomal clondronate in vivo. Fundamental experiments and functional assays, hematoxylin and eosin staining, oil red O staining and others, were performed to evaluate the function of AI662270 on atherogenesis. Peritoneal macrophages were treated with oxidized low density lipoprotein (ox-LDL) to simulate in vitro model. Mechanism assays, RNA-interacting protein immunoprecipitation, RNA–protein pulldown and others, were performed to study the regulatory mechanism of AI662270 in macrophages. Results The novel AI662270 was mainly enriched in macrophages, but not in endothelial cells, smooth muscle cells and fibroblasts of mouse atherosclerotic lesions and was upregulated by ox-LDL. Overexpression of AI662270 resulted in lipid accumulation, larger atherosclerotic plaques and cardiac dysfunction in vivo. After macrophages were removed, the pro-atherogenic effect of AI662270 disappeared. Downregulation of AI662270 in macrophages protected against foam cell formation by potentiating cholesterol efflux and reducing intracellular total cholesterol. The opposite effect was observed in macrophage-specific AI662270-overexpressed cells in vitro. AI662270 bound to adenosine triphosphate-binding cassette transporter A1 (Abca1) responsible for regulating cholesterol efflux in macrophages. Forced expression of AI662270 in macrophages decreased Abca1 expression. The reverse occurred when expression of AI662270 was repressed. Conclusion These findings reveal an essential role for AI662270 in atherosclerosis progression by regulating cholesterol efflux from macrophages.

Atherosclerosis is a complex inflammatory disease that affects the arteries and can lead to severe complications such as heart attack and stroke. Macrophages, a type of immune cell, play a crucial role in atherosclerosis initiation and progression. Emerging studies revealed that ion channels regulate macrophage activation, polarization, phagocytosis, and cytokine secretion. Moreover, macrophage ion channel dysfunction is implicated in macrophage-derived foam cell formation and atherogenesis. In this context, exploring the regulatory role of ion channels in macrophage function and their impacts on the progression of atherosclerosis emerges as a promising avenue for research. Studies in the field will provide insights into novel therapeutic targets for the treatment of atherosclerosis. [ABSTRACT FROM AUTHOR]

Aims: Atherosclerosis is a huge threaten to the human health, C1q/TNF‐related protein 9 (CTRP9) has been previously reported possessing vascular protective functions. Our study is aimed to reveal the mechanism of the regulative effects of CTRP9 on the foam cell formation. Methods and results: Primary human macrophages were isolated from human monocytes donated by healthy volunteers. CCK‐8 assay was performed for determining the cell viability. Oil Red O staining was employed for measuring the lipid accumulation. Cholesterol ester and cholesterol concentration were detected by commercial kits for evaluating the intracellular cholesterol. Ubiquitination assay was performed to reveal the ubiquitination level of CD36, cycloheximide assay was applied for determining the half‐life of CD36 protein. Quantitative real‐time PCR and western blot assays were performed for detecting the mRNA and protein expression. Pre‐treatment with CTRP9 in primary human macrophages markedly suppressed the cholesterol accumulation concentration after oxidized low‐density lipoprotein treatment. CD36 was significantly increased after oxidized low‐density lipoprotein exposure while was reduced by CTRP9 treatment. Up‐regulation of CD36 significantly reversed the CTRP9‐mediated protective effects in foam cells. The differential expression levels of several deubiquitinating enzymes preliminarily indicated that USP11 was obviously decreased after CTRP9 treatment. USP11 knockdown decreased the CD36 protein expression and pre‐treatment with 10 μg/mL MG132 significantly maintained the CD36 level from USP11 knock down. Up‐regulation of CD36 reversed the alterations on the cholesterol metabolism caused by CTRP9 or USP11 knockdown. Conclusions: CTRP9 regulates the USP11/CD36 axis to protect the macrophages form transforming into foam cells by suppressing intracellular lipid and cholesterol accumulation, which is a potential therapeutic agent for atherosclerosis. [ABSTRACT FROM AUTHOR]

To explore the sesquiterpenoids in Curcuma longa L. and their activity related to anti-atherosclerosis. The chemical compounds of the rhizomes of C. longa were separated and purified by multiple chromatography techniques. Their structures were established by a variety of spectroscopic experiments. The absolute configurations were determined by comparing experimental and calculated NMR chemical shifts and electronic circular dichroism (ECD) spectra. Their anti-inflammatory effects and inhibitory activity against macrophage-derived foam cell formation were evaluated by lipopolysaccharide (LPS) and oxidized low-density lipoprotein (ox-LDL)-injured RAW264.7 macrophages, respectively. This study resulted in the isolation of 10 bisabolane-type sesquiterpenoids (1–10) from C. longa, including two pairs of new epimers (curbisabolanones A–D, 1–4). Compound 4 significantly inhibited LPS-induced nitric oxide (NO), interleukin-1β (IL-1β), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), and prostaglandin E2 (PGE2) production in RAW264.7 cells. Furthermore, compound 4 showed inhibitory activity against macrophage-derived foam cell formation, which was represented by markedly reducing ox-LDL-induced intracellular lipid accumulation as well as total cholesterol (TC), free cholesterol (FC), and cholesterol ester (CE) contents in RAW264.7 cells. These findings suggest that bisabolane-type sesquiterpenoids, one of the main types of components in C. longa, have the potential to alleviate the atherosclerosis process by preventing inflammation and inhibiting macrophage foaming. [ABSTRACT FROM AUTHOR]

Atherosclerotic morbidity is significantly higher in the diabetic population. Hyperglycemia, a typical feature of diabetes, has been proven to accelerate foam cell formation. However, the molecular mechanisms behind this process remain unclear. In this study, LPS and IFN-γ were used to convert THP-1-derived macrophages into M1 macrophages, which were then activated with ox-LDL in either high glucose or normal condition. We identified lipids within macrophages by Oil red O staining and total cholesterol detection. The genes involved in lipid absorption, efflux, inflammation, and metabolism were analyzed using qRT-PCR. The mechanisms of high glucose-induced foam cell formation were further investigated through metabolomics and transcriptomics analysis. We discovered that high glucose speed up lipid accumulation in macrophages (both lipid droplets and total cholesterol increased), diminished lipid efflux (ABCG1 down-regulation), and aggravated inflammation (IL1B and TNF up-regulation). Following multi-omics analysis, it was determined that glucose altered the metabolic and transcriptional profiles of macrophages, identifying 392 differently expressed metabolites and 293 differentially expressed genes, respectively. Joint pathway analysis suggested that glucose predominantly disrupted the glycerolipid, glycerophospholipid, and arachidonic acid metabolic pathways in macrophages. High glucose in the glyceride metabolic pathway, for instance, suppressed the transcription of triglyceride hydrolase (LIPG and LPL), causing cells to deposit excess triglycerides into lipid droplets and encouraging foam cell formation. More importantly, high glucose triggered the accumulation of pro-atherosclerotic lipids (7-ketocholesterol, lysophosphatidylcholine, and glycerophosphatidylcholine). In conclusion, this work elucidated mechanisms of glucose-induced foam cell formation via a multi-omics approach. [ABSTRACT FROM AUTHOR]

Monocyte-to-macrophage differentiation and subsequent foam cell formation are key processes that contribute to plaque build-up during the progression of atherosclerotic lesions. Palmitoylation enzymes are known to play pivotal roles in the development and progression of inflammatory diseases. However, their specific impact on atherosclerosis development remains unclear. In this study, we discovered that the knockout of zDHHC1 in THP-1 cells, as well as Zdhhc1 in mice, markedly reduces the uptake of oxidized low-density lipoprotein (ox-LDL) by macrophages, thereby inhibiting foam cell formation. Moreover, the absence of Zdhhc1 in ApoE−/− mice significantly suppresses atherosclerotic plaque formation. Mass spectrometry coupled with bioinformatic analysis revealed an enrichment of the PI3K-Akt-mTOR signaling pathway. Consistent with this, we observed that knockout of zDHHC1 significantly decreases the palmitoylation levels of p110α, a crucial subunit of PI3K. Notably, the deletion of Zdhhc1 facilitates the nuclear translocation of p110α in macrophages, leading to a significant reduction in the downstream phosphorylation of Akt at Ser473 and mTOR at Ser2448. This cascade results in a decreased number of macrophages within plaques and ultimately mitigates the severity of atherosclerosis. These findings unveil a novel role for zDHHC1 in regulating foam cell formation and the progression of atherosclerosis, suggesting it as a promising target for clinical intervention in atherosclerosis therapy. • ZDHHC1 deficiency significantly reduces foam cell formation by inhibiting the uptake of ox-LDL; • The absence of Zdhhc1 in ApoE–/– mice markedly suppresses atherosclerotic plaque formation; • Zdhhc1 deficiency mitigates atherosclerosis by inhibiting PI3K-Akt-mTOR via promoting p110α nuclear translocation. [ABSTRACT FROM AUTHOR]

Background: Gastrodin is an effective polyphenol extracted from Chinese natural herbal Gastrodiae elata Blume, which exhibits antioxidant and anti-inflammatory effects. It has been reported to benefit neurodegenerative diseases, but the effect of Gastrodin on atherosclerosis and the underlying mechanisms remain elusive. The aim of this study is to investigate the function and mechanism of Gastrodin in atherosclerosis. Methods: Atherosclerosis mouse model was established by fed low density lipoprotein receptor-deficient (Ldlr−/−) mice with a high fat diet (HFD, 20% fat and 0.5 cholesterol) for 8 weeks and Gastrodin was administered daily via oral gavage. Plasma lipid levels were measured using commercial kits. En face and aortic sinus lipid accumulation were analyzed with Oil Red O staining. In vitro cell models using foam cell formation model and classical atherosclerosis inflammation model, macrophages were incubated with oxygenized low-density lipoproteins (ox-LDL) or lipopolysaccharide (LPS) in the presence of different concentration of Gastrodin or vehicle solution. Foam cell formation and cellular lipid content were evaluated by Oil Red O staining and intracellular lipids extraction analysis. Gene expression and proteins related to cholesterol influx and efflux were examined by quantitative reverse transcription PCR (RT-qPCR) and western blotting analysis. Furthermore, the effect of Gastrodin on LPS induced macrophage inflammatory responses and NF-κB pathway were evaluated by RT-qPCR and western blotting analysis. Results: Gastrodin administration reduced the body weight, plasma lipid levels in Ldlr−/− mice after fed a high fat diet. Oil Red O staining showed Gastrodin-treated mice displayed less atherosclerosis lesion area. Furthermore, Gastrodin treatment significantly ameliorated ox-LDL-induced macrophage-derived foam cells formation through suppressing genes expression related to cholesterol efflux including scavenger receptor class B and ATP-binding cassette transporter A1. Moreover, Gastrodin markedly suppressed pro-inflammatory cytokines secretion and LPS induced inflammatory response in macrophage through downregulating NF-κB pathway. Conclusions: Our study demonstrated that Gastrodin attenuates atherosclerosis by suppressing foam cells formation and LPS-induced inflammatory response and represents a novel therapeutic target for the treatment of atherosclerosis. [ABSTRACT FROM AUTHOR]

Background: Atherosclerosis is driven by synergistic interactions between pathological biomechanical and lipid metabolic factors. Long noncoding RNAs (LncRNAs) have been implicated in atherogenesis. The purpose of this study was to investigate the potential mechanism of lncRNA AI662270 on macrophage cholesterol transport in atherosclerosis. Methods: Apolipoprotein E deficiency (ApoE−/−) mice were fed a high fat diet for 16 weeks to construct atherosclerotic model, and the mice were injected with recombinant lentivirus carrying AI662270 gene to overexpress AI662270. Macrophages were cleared by liposomal clondronate in vivo. Fundamental experiments and functional assays, hematoxylin and eosin staining, oil red O staining and others, were performed to evaluate the function of AI662270 on atherogenesis. Peritoneal macrophages were treated with oxidized low density lipoprotein (ox-LDL) to simulate in vitro model. Mechanism assays, RNA-interacting protein immunoprecipitation, RNA–protein pulldown and others, were performed to study the regulatory mechanism of AI662270 in macrophages. Results: The novel AI662270 was mainly enriched in macrophages, but not in endothelial cells, smooth muscle cells and fibroblasts of mouse atherosclerotic lesions and was upregulated by ox-LDL. Overexpression of AI662270 resulted in lipid accumulation, larger atherosclerotic plaques and cardiac dysfunction in vivo. After macrophages were removed, the pro-atherogenic effect of AI662270 disappeared. Downregulation of AI662270 in macrophages protected against foam cell formation by potentiating cholesterol efflux and reducing intracellular total cholesterol. The opposite effect was observed in macrophage-specific AI662270-overexpressed cells in vitro. AI662270 bound to adenosine triphosphate-binding cassette transporter A1 (Abca1) responsible for regulating cholesterol efflux in macrophages. Forced expression of AI662270 in macrophages decreased Abca1 expression. The reverse occurred when expression of AI662270 was repressed. Conclusion: These findings reveal an essential role for AI662270 in atherosclerosis progression by regulating cholesterol efflux from macrophages. [ABSTRACT FROM AUTHOR]

Nonalcoholic steatohepatitis (NASH) is the progressive form of nonalcoholic fatty liver disease (NAFLD), and the dysregulation of lipid metabolism and oxidative stress are the typical features. Subsequent dyslipidemia and oxygen radical production may render the formation of modified lipids. Macrophage scavenger receptor 1 (MSR1) is responsible for the uptake of modified lipoprotein and is one of the key molecules in atherosclerosis. However, the unrestricted uptake of modified lipoproteins by MSR1 and the formation of cholesterol-rich foamy macrophages also can be observed in NASH patients and mouse models. In this review, we highlight the dysregulation of lipid metabolism and oxidative stress in NASH, the alteration of MSR1 expression in physiological and pathological conditions, the formation of modified lipoproteins, and the role of MSR1 on macrophage foaming and NASH development and progression. [ABSTRACT FROM AUTHOR]

The accumulation of oxidized low-density lipoprotein (oxLDL) in macrophages leads to the formation of foam cells and atherosclerosis development. Reducing the uptake of oxLDL in macrophages decreases the incidence and progression of atherosclerosis. Four distinct single-strand DNA sequences, namely, AP07, AP11, AP25, and AP29, were selected that demonstrated specific binding to distinct regions of oxidized apolipoprotein B100 (apoB100; the protein component of oxLDL) with low HDOCK scores. These four DNA sequences were combined to generate aptamers that selectively bound to labeled Dil-oxLDL, and were subsequently added to murine RAW 264.7 macrophages to test their inhibitory effects using fluorescence spectrometry. The four combined aptamers at 10 μM reduced oxLDL uptake by 79 ± 4% compared to that of the untreated aptamer group. Flow cytometry data demonstrated that macrophages treated with aptamers reached only 32.6% of the Dil-oxLDL signal, a 50% reduction in fluorescence emission relative to that of the untreated group (64.4% Dil-oxLDL signal). Binding the four combined aptamers to the oxLDL surface disrupted the interaction between oxLDL and CD36 via cyclic voltammetry, effectively decreasing the level of uptake of oxLDL by macrophages. Results suggested that these aptamers could be used as alternative compounds to prevent the formation of foam cells, hence providing antiatherosclerosis activity.

The aim of this study was to investigate the underlying mechanism of chrysophanol(Chr) in reducing inflammation and foam cell formation induced by oxidized low-density lipoprotein(ox-LDL) and to investigate the targets and pathways related to effects of Chr on coronary atherosclerosis, providing a theoretical basis for the development of new clinical drugs. RAW264.7 macrophages were cultured in vitro, and after determining the appropriate concentrations of Chr and ox-LDL for treating RAW264.7 macrophages using a cell counting kit-8(CCK-8), the macrophages were treated with different concentrations of Chr(10, 15 μmol·L~(-1)) and ox-LDL(with or without 80 mg·mL~(-1)) for 24 h. RAW264.7 macrophages were divided into four groups: control group, model group(80 mg·mL~(-1) ox-LDL), treatment group(80 mg·mL~(-1) ox-LDL+10 μmol·L~(-1) Chr), and treatment group(80 mg·mL~(-1) ox-LDL+15 μmol·L~(-1) Chr). Lipid accumulation in each group was detected by oil red O staining. CD36 expression was analyzed by flow cytometry. Western blot was used to detect the expression of scavenger receptor class A1(SR-A1), scavenger receptor class B type Ⅰ(SR-B1), autophagy-related protein 5(Atg5), Beclin-1, autophagy adaptor protein p62(P62), the ratio of microtubule-associated protein light chain 3(LC3)Ⅱ to LC3Ⅰ(LC3Ⅱ/LC3Ⅰ), nuclear factor kappa B P65(NF-κB P65), inhibitor of κB kinase β(IKKβ), nuclear factor of κB inhibitor(IκB), high mobility group box protein 1(HMGB1), phosphatidylinositol 3-kinase(PI3K), protein kinase B(Akt), and phosphorylated mammalian target of rapamycin(mTOR). Real-time quantitative polymerase chain reaction(RT-qPCR) was used to detect the mRNA expression levels of ATP-binding cassette transporter A1(ABCA1), ATP-binding cassette transporter G1(ABCG1), interleukin-1β(IL-1β), tumor necrosis factor-α(TNF-α), HMGB1, inducible nitric oxide synthase(iNOS), arginase 1(Arg1), macrophage galactose-type lectin-1(Mgl-1), and NF-κB P65. Immunofluorescence analysis was performed to determine the localization of HMGB1 in RAW264.7 cells in each group. The autophagy inhibitor 3-methyladenine(3-MA) was added as a control for reverse validation, and the RAW264.7 macrophages were divided into four groups again: control group, model group(80 mg·mL~(-1) ox-LDL), treatment group(80 mg·mL~(-1) ox-LDL + 15 μmol·L~(-1) Chr), and inhibitor group(80 mg·mL~(-1) ox-LDL+15 μmol·L~(-1) Chr+3-MA). The results showed that Chr effectively reduced foam cell formation by regulating the expression levels of SR-A1, ABCA1, ABCG1, the LC3Ⅱ/LC3Ⅰ ratio, Atg5, Beclin-1, and p62, and inhibited the NF-κB/HMGB1-PI3K/Akt/mTOR signaling pathway. Moreover, the inhibitory effects of Chr on autophagy and the NF-κB/HMGB1-PI3K/Akt/mTOR pathway were reversed by the autophagy inhibitor 3-MA. In conclusion, Chr exhibits therapeutic potential for the treatment of atherosclerosis by inducing autophagy and modulating the NF-κB/HMGB1 and PI3K/Akt/mTOR pathways to inhibit the formation of macrophage inflammatory foam cells.

Atherosclerosis (AS) is the main pathological basis for the development of cardiovascular diseases. Vascular inflammation is an important factor in the formation of AS, and macrophage pyroptosis plays a key role in AS due to its unique inflammatory response. Guizhitongluo Tablet (GZTLT) has shown clinically effective in treating patients with AS, but its mechanism is elusive. This study was to determine the effects of GZTLT on atherosclerotic vascular inflammation and pyroptosis and to understand its underlying mechanism. The active constituents of GZTLT were analysed by means of UPLC-HRMS. In vivo experiments were performed using ApoE−/− mice fed a high fat diet for 8 weeks, followed by treatment with varying concentrations of GZTLT orally by gavage and GsMTx4 (GS) intraperitoneally and followed for another 8 weeks. Oil red O, Haematoxylin-eosin (HE) and Masson staining were employed to examine the lipid content, plaque size, and collagen fibre content of the mouse aorta. Immunofluorescence staining was utilised to identify macrophage infiltration, as well as the expression of Piezo1 and NLRP3 proteins in aortic plaques. The levels of aortic inflammatory factors were determined using RT-PCR and ELISA. In vitro, foam cell formation in bone marrow-derived macrophages (BMDMs) was observed using Oil Red O staining. Intracellular Ca2+ measurements were performed to detect the calcium influx in BMDMs, and the expression of NLRP3 and its related proteins were detected by Western blot. The UPLC-HRMS analysis revealed 31 major components of GZTLT. Our data showed that GZTLT inhibited aortic plaque formation in mice and increased plaque collagen fibre content to stabilise plaques. In addition, GZTLT could restrain the expression of serum lipid levels and suppress macrophage foam cell formation. Further studies found that GZTLT inhibited macrophage infiltration in aortic plaques and suppressed the expression of inflammatory factors. It is noteworthy that GZTLT can restrain Piezo1 expression and reduce Ca2+ influx in BMDMs. Additionally, we found that GZTLT could regulate NLRP3 activation and pyroptosis by inhibiting Piezo1. The present study suggests that GZTLT inhibits vascular inflammation and macrophage pyroptosis through the Piezo1/NLRP3 signaling pathway, thereby delaying AS development. Our finding provides a potential target for AS treatment and drug discovery. [Display omitted] [ABSTRACT FROM AUTHOR]

Macrophages are key inflammatory immune cells that display dynamic phenotypes and functions in response to their local microenvironment. In different conditions, macrophage polarization can be induced by high-mobility group box 1 (HMGB1), a nuclear DNA-binding protein that activates innate immunity via the Toll-like receptor (TLR) 4, the receptor for advanced glycation end products (RAGE), and C-X-C chemokine receptor (CXCR) 4. This study investigated the phenotypes of murine bone-marrow-derived macrophages (BMDMs) stimulated with different HMGB1 redox isoforms using bulk RNA sequencing (RNA-Seq). Disulfide HMGB1 (dsHMGB1)-stimulated BMDMs showed a similar but distinct transcriptomic profile to LPS/IFNγ- and LPS-stimulated BMDMs. Fully reduced HMGB1 (frHMGB1) did not induce any significant transcriptomic change. Interestingly, compared to LPS/IFNγ- and LPS-, dsHMGB1-stimulated BMDMs showed lipid metabolism and foam cell differentiation gene set enrichment, and oil red O staining revealed that both dsHMGB1 and frHMGB1 alleviated oxidized low-density lipoprotein (oxLDL)-induced foam cells formation. Overall, this work, for the first time, used transcriptomic analysis by RNA-Seq to investigate the impact of HMGB1 stimulation on BMDM polarization. Our results demonstrated that dsHMGB1 and frHMGB1 induced distinct BMDM polarization phenotypes compared to LPS/IFNγ- and LPS- induced phenotypes. [ABSTRACT FROM AUTHOR]

Abstract Background Premature atherosclerosis occurs in patients with SLE; however, the mechanisms remain unclear. Both mitochondrial machinery and proinflammatory cytokine interferon alpha (IFN-α) potentially contribute to atherogenic processes in SLE. Here, we explore the roles of the mitochondrial protein cytidine/uridine monophosphate kinase 2 (CMPK2) in IFN-α-mediated pro-atherogenic events. Methods Foam cell measurements were performed by oil red O staining, Dil-oxLDL uptake and the BODIPY approach. The mRNA and protein levels were measured by qPCR and Western blotting, respectively. Isolation of CD4+ T cells and monocytes was performed with monoclonal antibodies conjugated with microbeads. Manipulation of protein expression was conducted by either small interference RNA (siRNA) knockdown or CRISPR/Cas9 knockout. The expression of mitochondrial reactive oxygen species (mtROS) was determined by flow cytometry and confocal microscopy. Results IFN-α enhanced oxLDL-induced foam cell formation and Dil-oxLDL uptake by macrophages. In addition to IFN-α, several triggers of atherosclerosis, including thrombin and IFN-γ, can induce CMPK2 expression, which was elevated in CD4+ T cells and CD14+ monocytes isolated from SLE patients compared to those isolated from controls. The analysis of cellular subfractions revealed that CMPK2 was present in both mitochondrial and cytosolic fractions. IFN-α-induced CMPK2 expression was inhibited by Janus kinase (JAK)1/2 and tyrosine kinase 2 (Tyk2) inhibitors. Both the knockdown and knockout of CMPK2 attenuated IFN-α-mediated foam cell formation, which involved the reduction of scavenger receptor class A (SR-A) expression. CMPK2 also regulated IFN-α-enhanced mtROS production and inflammasome activation. Conclusions The study suggests that CMPK2 plays contributing roles in the pro-atherogenic effects of IFN-α.

Atherosclerosis, accompanied by inflammation and metabolic disorders, is the primary cause of clinical cardiovascular death. In recent years, unhealthy lifestyles (e.g., sedentary lifestyles) have contributed to a worldwide epidemic of atherosclerosis. Exercise is a known treatment of atherosclerosis, but the precise mechanisms are still unknown. Here, we show that 12 weeks of regular exercise training on a treadmill significantly decreased lipid accumulation and foam cell formation in ApoE−/− mice fed with a Western diet, which plays a critical role in the process of atherosclerosis. This was associated with an increase in β-hydroxybutyric acid (BHB) levels in the serum. We provide evidence that BHB treatment in vivo or in vitro increases the protein levels of cholesterol transporters, including ABCA1, ABCG1, and SR-BI, and is capable of reducing lipid accumulation. It also ameliorated autophagy in macrophages and atherosclerosis plaques, which play an important role in the step of cholesterol efflux. Altogether, an increase in serum BHB levels after regular exercise is an important mechanism of exercise inhibiting the development of atherosclerosis. This provides a novel treatment for atherosclerotic patients who are unable to undertake regular exercise for whatever reason. They will gain a benefit from receiving additional BHB. [ABSTRACT FROM AUTHOR]

Background: The production of lipid-laden cells in macrophages after significant ingestion of oxidized low-density lipoprotein is considered the most critical phase in the creation of atherosclerotic lesions, which is known as foam cell formation. Targeting foam cell development to find a potential therapeutic strategy for the management of atherosclerosis has yielded numerous promising outcomes. Multiple variables influence foam cell growth, including scavenger receptor expression, cholesterol transporter expression acyl CoA: cholesterol acyltransferase activity, and neutral cholesteryl ester hydrolase activity. Plants used during herbal therapy have been shown to assist with a variety of ailments. Result: In this study, we found medicinal plants and their bioactive components suppress foam cell formation in a variety of ways; some inhibit cholesterol transporter and lectin-like oxidized low-density lipoprotein receptor-1 upregulation, while others inhibit the function of acyl CoA: cholesterol acyltransferase activity, and neutral cholesteryl ester hydrolase activity. Conclusion: Recent study findings related to the synthesis of the new active component from plant sources by focusing on the typical process involved in the generation of foam cells. We're also looking at using a cellular target-based therapeutic approach to generate novel plant-based medications for the cure of atherosclerosis. [ABSTRACT FROM AUTHOR]

Abstract Background Macrophages that accumulate in atherosclerotic plaques contribute to progression of the lesions to more advanced and complex plaques. Although iron deposition was found in human atherosclerotic plaques, clinical and pre-clinical studies showed controversial results. Several epidemiological studies did not show the positive correlation between a systemic iron status and an incidence of cardiovascular diseases, suggesting that the iron involvement occurs locally, rather than systemically. Results To determine the direct in vivo effect of iron accumulation in macrophages on the progression of atherosclerosis, we generated Apoe −/− mice with a macrophage-specific ferroportin (Fpn1) deficiency (Apoe −/− Fpn1 LysM/LysM ). Fpn1 deficiency in macrophages dramatically accelerated the progression of atherosclerosis in mice. Pathophysiological evidence showed elevated levels of reactive oxygen species, aggravated systemic inflammation, and altered plaque-lipid composition. Moreover, Fpn1 deficiency in macrophages significantly inhibited the expression of ABC transporters (ABCA1 and ABCG1) by decreasing the expression of the transcription factor LXRα, which reduced cholesterol efflux and therefore promoted foam cell formation and enhanced plaque formation. Iron chelation relieved the symptoms moderately in vivo, but drastically ex vivo. Conclusions Macrophage iron content in plaques is a critical factor in progression of atherosclerosis. The interaction of iron and lipid metabolism takes place in macrophage-rich atherosclerotic plaques. And we also suggest that altering intracellular iron levels in macrophages by systemic iron chelation or dietary iron restriction may be a potential supplementary strategy to limit or even regress the progression of atherosclerosis.

Macrophages are critical components of atherosclerotic lesions and their pro- and anti-inflammatory responses influence atherogenesis. Type-I interferons (IFNs) are cytokines that play an essential role in antiviral responses and inflammatory activation and have been shown to promote atherosclerosis. Although the impact of type-I IFNs on macrophage foam cell formation is well-documented, the effect of lipid accumulation in monocytes and macrophages on type-I IFN responses remains unknown. Here we examined IFN stimulated (ISG) and non-ISG inflammatory gene expression in mouse and human macrophages that were loaded with acetylated LDL (acLDL), as a model for foam cell formation. We found that acLDL loading in mouse and human macrophages specifically suppressed expression of ISGs and IFN-β secretion, but not other pro-inflammatory genes. The down regulation of ISGs could be rescued by exogenous IFN-β supplementation. Activation of the cholesterol-sensing nuclear liver X receptor (LXR) recapitulated the cholesterol-initiated type-I IFN suppression. Additional analyses of murine in vitro and in vivo generated foam cells confirmed the suppressed IFN signaling pathways and suggest that this phenotype is mediated via down regulation of interferon regulatory factor binding at gene promoters. Finally, RNA-seq analysis of monocytes of familial hypercholesterolemia (FH) patients also showed type-I IFN suppression which was restored by lipid-lowering therapy and not present in monocytes of healthy donors. Taken together, we define type-I IFN suppression as an athero-protective characteristic of foamy macrophages. These data provide new insights into the mechanisms that control inflammatory responses in hyperlipidaemic settings and can support future therapeutic approaches focusing on reprogramming of macrophages to reduce atherosclerotic plaque progression and improve stability.

Protein phosphatase 2A (PP2A), a crucial serine/threonine phosphatase, has recently been reported to play an important role in cardiovascular disease. Previous studies have hinted that PP2A is involved in atherosclerosis formation, but the associated mechanisms remain poorly understood. In this study, we investigate the role of PP2A in the pathogenesis of atherosclerosis. In human atherosclerotic coronary arteries, we found that the expression and activity of PP2A decreased significantly when compared to non-atherosclerotic arteries. Additional experiments demonstrated that pharmacological inhibition of PP2A aggravated atherosclerosis of ApoE−/− mice. Considering the central role of macrophages in atherosclerosis, mice with conditional knockout of the PP2A-Cα subunit in myeloid cells were produced to investigate the function of PP2A in macrophages. Results showed that PP2A deficiency in myeloid cells aggravated atherosclerotic lesions in mice. in vitro experiments indicated that PP2A-deficient macrophages had an enhanced ability of lipid uptake and foam cell formation. Mechanistically, the deficiency of the PP2A in macrophages led to an increase in the phosphorylation level of p38, which contributed to the elevated expression of scavenger receptor CD36, a key factor involved in lipoprotein uptake. Our data suggest that PP2A participates in the pathophysiological process of atherosclerosis. The decrease of PP2A expression and activity in macrophages is a crucial determinant for foam cell formation and the initiation of atherosclerosis. Our study may provide a potential novel approach for the treatment of atherosclerosis.

Deciphering the molecular and cellular processes involved in foam cell formation is critical for us to understand the pathogenesis of atherosclerosis. Nuclear factor of activated T cells (NFAT) is a transcription factor originally identified as a key player in the differentiation of T cells and maturation of immune system. Nowadays it has been brought into attention that NFAT also regulates multiple pathophysiological processes and targeted intervention in NFAT may be effective in the treatment of some cardiovascular diseases. However, whether NFAT is involved in foam cell formation remains elusive. NFAT in human monocyte‐derived macrophage was activated by ox‐LDL and translocated from the cytoplasm to the nucleus. NFAT then directly bound to peroxisome proliferator‐activated receptor γ (PPARγ) in the nucleus and negatively regulated its transcriptional activity. NFATc2 knockdown or NFAT inhibitor 11R‐VIVIT increased cholesterol efflux (by activating PPARγ‐LXRα‐ABCA1 cascade) and reduced the uptake of modified lipoprotein (in a PPARγ‐independent way) in macrophage, thus prevented foam cell formation. Besides, 11R‐VIVIT also exerted a protective role in the development of atherosclerosis in western diet‐fed ApoE−/− mice. These results suggest NFAT inhibition as a potential therapeutic strategy in atherosclerosis. [ABSTRACT FROM AUTHOR]

• Angiopoietin-like proteins are involved in atherosclerosis-related pathological processes. • Angiopoietin-like proteins can be the roles as independent predictive value of biomarkers for atherosclerosis therapy. • The precise mechanism that Angiopoietin-like proteins regulate atherosclerosis process deserve to be further explored. Atherosclerosis, as a chronic inflammatory disease within the arterial wall, is a leading cause of morbidity and mortality worldwide due to its role in myocardial infarction, stroke and peripheral artery disease. Additional evidence is emerging that the angiopoietin-like (ANGPTL) family of proteins participate in the pathology of this disease process via endothelial dysfunction, inflammation, dyslipidemia, calcification, foam cell formation and platelet activation. This review summarizes current knowledge on the ANGPTL family of proteins in atherosclerosis related pathological processes. Moreover, the potential value of ANGPTL family proteins as predictive biomarkers in atherosclerosis is discussed. Given the attractive role of ANGPTL3, ANGPTL4, ANGPTL8 in atherosclerotic dyslipidemia via regulation of lipoprotein lipase (LPL), antisense oligonucleotide or/and monoclonal antibody-based inactivation of these proteins represent potential atherosclerotic therapies. [ABSTRACT FROM AUTHOR]

Huanglian Jiedu Decoction (HLJDD) is a Chinese medicine with a long history of therapeutic application. It is widely used in treating atherosclerosis (AS) in Chinese medicine theory and clinical practice. However, the mechanism of HLJDD in treating AS remains unclear. To investigate the efficacy and mechanism of HLJDD in treating AS. AS was induced on high-fat diet-fed ApoE−/− mice, with the aorta pathological changes evaluated with lipid content and plaque progression. In vitro, foam cells were induced by subjecting primary mouse aortic vascular smooth muscle cells (VSMCs) to oxLDL incubation. After HLJDD intervention, VSMCs were assessed with lipid stack, apoptosis, oxidative stress, and the expression of foam cell markers. The effects of P2RY12 were tested by adopting clopidogrel hydrogen sulfate (CDL) in vivo and transfecting P2RY12 over-expressive plasmid in vitro. Autophagy was inhibited by Chloroquine or transfecting siRNA targeting ATG7 (siATG7). The mechanism of HLJDD treating atherosclerosis was explored using network pharmacology and validated with molecular docking and co-immunoprecipitation. HLJDD exhibited a dose-dependent reduction in lipid deposition, collagen loss, and necrosis within plaques. It also reversed lipid accumulation and down-regulated the expression of foam cell markers. P2RY12 inhibition alleviated AS, while P2RY12 overexpression enhanced foam cell formation and blocked the therapeutic effects of HLJDD. Network pharmacological analysis suggested that HLJDD might mediate PI3K/AKT signaling pathway-induced autophagy. P2RY12 overexpression also impaired autophagy. Similarly, inhibiting autophagy counteracted the effect of CDL, exacerbated AS in vivo, and promoted foam cell formation in vitro. However, HLJDD treatment mitigated these detrimental effects by suppressing the PI3K/AKT signaling pathway. Immunofluorescence and molecular docking revealed a high affinity between P2RY12 and PIK3CB, while co-immunoprecipitation assays illustrated their interaction. HLJDD inhibited AS in vivo and foam cell formation in vitro by restoring P2RY12/PI3K/AKT signaling pathway-suppressed autophagy. This study is the first to reveal an interaction between P2RY12 and PI3K3CB. [Display omitted] • Huanglian Jiedu Decoction (HLJDD) inhibited high-fat diet-induced atherosclerosis (AS). • HLJDD suppressed vascular smooth muscle cell-derived foam cell formation. • HLJDD restored P2RY12/PI3K/AKT signaling pathway-suppressed autophagy in AS. • P2RY12 and PIK3CB formed a protein complex in immunoprecipitation. [ABSTRACT FROM AUTHOR]

Vascular smooth muscle cells (SMCs) are important vascular components that are essential for the regulation of vascular functions during vascular atherosclerogenesis and vascular injury. Oxidized low-density lipoprotein (oxLDL) is known to induce SMC activation and foam cell transformation. This study characterized the role of hepatoma-derived growth factor (HDGF) in oxLDL-induced foam cell formation in cultured primary rat aortic SMCs. OxLDL exposure significantly increased HDGF expression and extracellular release. It also upregulated atherogenic regulators in SMCs, including TLR4, MyD88, LOX-1, and CD36. Exogenous HDGF stimulation not only increased the expression of cognate receptor nucleolin, but also the innate immunity regulators TLR4/MyD88 and lipid metabolism regulators, including LOX-1 and CD36. Oil red O staining showed that HDGF did not initiate, but enhanced oxLDL-driven foam cell formation in SMCs. Further signaling characterization demonstrated that oxLDL evoked activation of PI3K/Akt and p38 MAPK signaling pathways, both of which were involved in the upregulation of HDGF, LOX-1, and CD36 induced by oxLDL. Gene knockdown experiments using LOX-1 targeted siRNA demonstrated that LOX-1 expression was critical for oxLDL-induced HDGF upregulation, while HDGF gene depletion completely abolished oxLDL-triggered TLR4, LOX-1, and CD36 overexpression and foam cell formation in SMCs. These findings strongly suggest that oxLDL-induced HDGF upregulation participates in subsequent LOX-1 and CD36 expression in aortic SMCs and mechanistically contributes to the formation of SMC-derived foam cells. The oxLDL/LOX-1/HDGF axis may serve as a target for anti-atherogenesis therapy.

Oleacein is a polyphenol and can be found in unfiltered extra virgin olive oil and leaves Ligustrum vulgare L. (Oleaceae). The aim of the present study was to investigate whether oleacein could increase efflux cholesterol from macrophages.Oleacein in dose-dependent manner (OLEA20, OLEA50) significantly reduced lipid deposits in macrophages. The increase of SRB1 expression was found for OLEA50 and OLEA20 compared to oxLDL-induced macrophages. Oleacein significantly up-regulated protein expression of ABCA1 and ABCG1 transporter. Moreover, oleacein increased of HO-1 intracellular secretion, as well as increased activation of Nrf2. Whereas, the level of NF-κB protein was almost completely inhibited by oleacein at the same concentrations. For OLEA10, no significant changes were observed for the parameters tested.Our study demonstrated that oleacein may reduce foam cell formation. Therefore, we propose a novel explanation for the oleacein, that can be used to develop new therapeutic strategies for the treatment of atherosclerosis.

Foam cell formation and inflammation in macrophages contribute to the development of atherosclerosis (AS). Clematichinenoside AR (AR) is a major active ingredient extracted from the traditional Chinese herb Clematis chinensis and has potent pharmacological effects on various diseases, including AS. However, little is known about the exact role and mechanism of AR in AS. RAW264.7 macrophages were exposed to oxidized low-density lipoprotein (ox-LDL) to induce AS in vitro. Cell viability was assessed by the CCK-8 assay. Foam cell formation was detected by Oil Red staining. Cholesterol levels were determined by corresponding commercial kits. The expression of inflammatory cytokines was detected by ELISA. Western blot and immunofluorescence assays were employed to detect the expression of corresponding genes. The results indicated that AR treatment inhibited the formation of foam cells and cholesterol accumulation but promoted cholesterol efflux by upregulating ABCA1/ABCG1 in ox-LDL-induced RAW264.7 macrophages. In addition, AR decreased the production of inflammatory cytokines by blunting the activation of the NLRP3 inflammasome and inducing autophagy. However, these effects of AR were weakened by the autophagy inhibitor bafilomycin A1 but were similar to those produced by the autophagy activator rapamycin. Collectively, our study provides novel insights into the beneficial effects of AR on promoting cholesterol efflux as well as inhibiting foam cell formation and inflammation by regulating autophagy, thus identifying AR as a promising therapeutic agent for the treatment of AS. [ABSTRACT FROM AUTHOR]

The persistence of macrophage-derived foam cells in the artery wall fuels atherosclerosis development. However, the mechanism of foam cell formation regulation remains elusive. We are committed to determining the role that CD147 might play in macrophage foam cell formation during atherosclerosis. In this study, we found that CD147 expression was primarily increased in mouse and human atherosclerotic lesions that were rich in macrophages and could be upregulated by ox-LDL. High-throughput compound screening indicated that ox-LDL-induced CD147 upregulation in macrophages was achieved through PI3K/Akt/mTOR signaling. Genetic deletion of macrophage CD147 protected against foam cell formation by impeding cholesterol uptake, probably through the scavenger receptor CD36. The opposite effect was observed in primary macrophages isolated from macrophage-specific CD147-overexpressing mice. Moreover, bioinformatics results indicated that CD147 suppression might exert an atheroprotective effect via various processes, such as cholesterol biosynthetic and metabolic processes, LDL and plasma lipoprotein clearance, and decreased platelet aggregation and collagen degradation. Our findings identify CD147 as a potential target for prevention and treatment of atherosclerosis in the future.

Gynostemma pentaphyllum was widely consumed in China because of its various pharmacological activities. Numerous studies have confirmed the beneficial effects of G. pentaphyllum. However, the ultimate compounds responsible for its inhibitory effects on foam cell formation were still unknown. Gypenoside LVI (GPLVI), a dammarane-type gynosaponin was isolated and identified from G. pentaphyllum. Treatment of GPLVI successfully attenuated foam cell formation, as evidenced by Oil Red O staining, TG and cholesterol contents, and Dil-ox-LDL uptake analyses. GPLVI displayed significant inhibitory effects on secretion of IL-6 and IL-1β. In contrast, production of TNF-α was not markedly reduced. GPLVI also markedly induced ABCG1, SRB1 and LXRα expression. Additionally, GPLVI significantly blocked phosphorylation of ERK and JNK. Taken together, our results suggested that GPLVI might be responsible for the inhibitory effect of G. pentaphyllum on ox-LDL-induced foam cell formation mainly by promoting cholesterol efflux and inhibiting inflammatory response.

Background/Aims: High mobility group B-1 (HMGB-1)-induced endoplasmic reticulum stress (ERS) has been implicated in inflammation and dendritic cell maturation. C/EBP-homologous protein (CHOP) is a vital component of ERS and apoptosis and plays a critical role in atherosclerosis. However, only a little information is available about the role of HMGB-1 in foam cell formation. Thus, the role of HMGB-1-induced ERS/CHOP pathway in apoptosis and formation of macrophage-derived foam cells is investigated. Methods: RAW264.7 cells were treated with oxidized low-density lipoprotein (oxLDL) in the absence and/or presence of HMGB-1, N-acetylcysteine (NAC, an antioxidant), glycyrrhizin (Gly, an HMGB-1 inhibitor), tunicamycin (TM, an ERS inducer), and 4-phenylbutyrate (4-PBA, an ERS inhibitor). Reactive oxygen species (ROS) production was examined by dihydroethidium (DHE) staining. Oil Red O staining, intracellular total cholesterol assay, and Dil-oxLDL uptake assay evaluated the accumulation of lipids in macrophages. Cell apoptosis was measured by flow cytometry and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. Western blot detected the expression of HMGB-1/ERS/CHOP pathway. Results: oxLDL induced HMGB-1 translocation and secretion in a dose- and time-dependent manner, which was inhibited by NAC. oxLDL-induced lipid accumulation in macrophages was promoted synergistically by HMGB-1 that was attenuated by Gly. Moreover, TM synergized with oxLDL induced lipid accumulation and apoptosis of macrophages; however, 4-PBA alleviated the oxLDL-induced apoptotic foam cells. Additionally, the inhibition of ERS with 4-PBA suppressed the expression of HMGB-1-induced CHOP. Conclusions: OxLDL triggered HMGB-1 secretion in macrophages via oxidative stress. Furthermore, HMGB-1 promoted the formation and apoptosis of macrophage-derived foam cells via activation of ERS/CHOP pathway.

Macrophage foam cell formation and inflammation are a pathological hallmark of atherosclerosis. ClC-2 has been implicated in various pathological processes, including inflammation and lipid metabolic disorder. However, the functional role of ClC-2 in macrophage foam cell formation and inflammation is unclear. Here, we found that ClC-2 was dominantly expressed in macrophages of atherosclerotic plaque and increased in atherogenesis. Knockdown of ClC-2 inhibited ox-LDL -induced lipid uptake and deposition in macrophages. The increase in CD36 expression and the decrease in ABCA1 expression induced by ox-LDL were alleviated by ClC-2 downregulation. Further, ClC-2 lacking limited the ox-LDL-induced secretion of inflammatory cytokines and chemokine, and suppressed Nlrp3 inflammasome activation. Restoration of Nlrp3 expression reversed the effect of ClC-2 downregulation on macrophage lipid accumulation and inflammation. Collectively, our study demonstrates that ClC-2 knockdown ameliorates ox-LDL-induced macrophage foam cell formation and inflammation by inhibiting Nlrp3 inflammasome activation. Macrophage ClC-2 lacking limits inflammation and foam cell formation, suggesting that ClC-2 may be a therapeutic target for atherosclerosis treatment. [ABSTRACT FROM AUTHOR]

Angiopoietin-like protein 8 (ANGPTL8) is a hormone involved in regulating lipid metabolism. Patients with coronary artery disease have markedly higher plasma levels of ANGPTL8 than controls; however, the role of ANGPTL8 in atherosclerosis has not been explored. Therefore, we explored the effects of ANGPTL8 on atherosclerosis development in a mouse model. We induced experimental atherosclerosis in ApoE −/− mice ANGPTL8 -knockdown. and ANGPTL8 -overexpression ApoE −/− mice. We also explored the mechanism using ANGPTL8 -overexpression macrophages. ANGPTL8 expression was increased in human and mouse atherosclerotic lesions. ANGPTL8 overexpression promoted the development of atherosclerosis whereas ANGPTL8 knockdown protected against atherosclerosis. Immunofluorescence co-staining results showed that ANGPTL8 was expressed in macrophages in atherosclerotic plaques. Compared with wild type cells, ANGPTL8 -overexpressing macrophages, including bone marrow-derived macrophages and Raw 264.7 macrophages, showed enhanced foam cell formation and increased accumulation of cholesterol that was induced by increased uptake and decreased efflux of cholesterol. The results of this study also showed that ANGPTL8 induced the expression of CD36 and scavenger receptor (SR)-A, and inhibited the expression of SR-BI. Our findings demonstrate an unanticipated role of ANGPTL8 in the development of atherosclerosis and regulation of foam cell formation. ANGPTL8 may be a promising new target for atherosclerosis. • ANGPTL8 promotes the development of atherosclerosis. • ANGPTL8 overexpression promotes macrophages to become foam cell that accumulate more cholesterol. • ANGPTL8 increases uptake and decreases efflux of cholesterol in macrophages. • ANGPTL8 induces the expression of CD36 and SR-A, and inhibits the expression of SR-BI. [ABSTRACT FROM AUTHOR]

(1) Background: The cholinergic anti-inflammatory pathway (CAP) has been implicated in the regulation of various diseases, including chronic inflammatory cardiovascular disorders such as atherosclerosis (AS). This study aims to explore the underlying regulatory mechanisms of CAP activity in the progression of AS. (2) Methods: The Apoe-/- mice were subjected to sham, bilateral cervical vagotomy surgery (VGX), and VGX supplemented with Gainesville Tokushima scientists (GTS)-21 (4 mg/kg/d) and then fed with a high-fat diet for 10 weeks. Atherosclerotic lesion size and inflammation levels were investigated by histology and inflammatory cytokines analysis. The blood M1/M2 macrophages were analyzed by flow cytometry. Primary mouse bone marrow-derived macrophages (BMDM), peritoneal macrophages, and RAW264.7 cells were treated with CAP agonists acetylcholine (Ach) and GTS-21 to study their effects on macrophage functions. (3) Results: Compared with the sham group, inhibition of CAP by the VGX resulted in growing aortic lipid plaque area, deteriorated inflammatory levels, and aberrant quantity of M1/M2 macrophages in Apoe-/- mice. However, these detrimental effects of VGX were significantly ameliorated by the reactivation of CAP through GTS-21 treatment. The in vitro study using macrophages revealed that stimulation with CAP agonists suppressed M1, but promoted M2 macrophage polarization through the upregulation of TNFAIP3 and phosphorylation STAT3 levels, respectively. Moreover, the activation of CAP inhibited the formation of macrophage foam cells in the peritoneal cavity by regulating genes related to cholesterol metabolism. (4) Conclusions: This study provides novel evidence and mechanisms that the CAP plays an important role in the regulation of AS development by controlling macrophage functions, implying a potential use of CAP activation as a therapeutic strategy for AS treatment.

Gynostemma pentaphyllum (Thunb.) Makino was traditionally used as antiatherogenic foods in China but ingredients responsible for the effects remained unexplored. Here, kaempferol 3-O-[2G-(E)-Coumaroyl-3G-O-β-D-glucosyl-3R-O-β-D-glucosylrutinoside] (KCGG), gypenoside XLVI (GPXLVI), gypenoside LI (GPLI), gypenoside L (GPL), ginsenoside Rd (GSRd) and 2α, 3β, 12β, 20 (S)-tetrahydroxy-25-hydroperoxy-dammar-23-ene-20-O-β-D-glucopyranosyl-(1 → 6)-β-D-glucopyranoside (GLUP) were isolated, among which KCGG, GPLI and GLUP exhibited greater antioxidant potential. The predominant inhibitory effects of KCGG, GPLI and GLUP on oxidized low-density lipoproteins (ox-LDL)-induced foam cell formation and intracellular lipid accumulation were further confirmed at non-cytotoxic concentrations. KCGG mainly decreased interleukin-6 (IL-6) production, while GPLI and GLUP were more powerful in inhibiting interleukin-1β (IL-1β) accumulation. KCGG, GPLI and GLUP significantly inhibited extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) phosphorylation. They also dramatically promoted cholesterol efflux through up-regulating scavenger receptor class B type I (SRB1) expression. In conclusion, KCGG, GPLI and GLUP acted through different pathways that acted simultaneously.

Oxidation of low-density lipoprotein (LDL) plays a crucial role in the pathogenesis of atherosclerosis. Hemin (iron (III)-protoporphyrin IX) is a degradation product of hemoglobin that can be found in thalassemia patients. Hemin is a strong oxidant that can cause LDL oxidation and contributes to atherosclerosis in thalassemia patients. Lusianthridin from Dendrobium venustrum is a phenolic compound that possesses antioxidant activity. Hence, lusianthridin could be a promising compound to be used against hemin-induced oxidative stress. The major goal of this study is to evaluate the protective effect of lusianthridin on hemin-induced low-density lipoprotein oxidation (he-oxLDL). Here, various concentrations of lusianthridin (0.25, 0.5, 1, and 2 µM) were preincubated with LDL for 30 min, then 5 µM of hemin was added to initiate the oxidation, and oxidative parameters were measured at various times of incubation (0, 1, 3, 6, 12, 24 h). Lipid peroxidation of LDL was measured by thiobarbituric reactive substance (TBARs) assay and relative electrophoretic mobility (REM). The lipid composition of LDL was analyzed by using reverse-phase HPLC. Foam cell formation with he-oxLDL in RAW 264.7 macrophage cells was detected by Oil Red O staining. The results indicated that lusianthridin could inhibit TBARs formation, decrease REM, decrease oxidized lipid products, as well as preserve the level of cholesteryl arachidonate and cholesteryl linoleate. Moreover, He-oxLDL incubated with lusianthridin for 24 h can reduce the foam cell formation in RAW 264.7 macrophage cells. Taken together, lusianthridin could be a potential agent to be used to prevent atherosclerosis in thalassemia patients.

Interleukin-17A (IL-17A), a major effector cytokine secreted by T helper 17 (Th17) cells, is elevated in atherosclerosis lesions. The purpose of this study was to assess the role of IL-17A in the pathogenesis of atherosclerosis. To measure the expression of adhesion molecules, human umbilical vein endothelial cells (HUVECs) and U937 cells were stimulated with IL-17A. Western blot and real-time polymerase chain reaction analyses revealed that intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) expression in HUVECs, and very late antigen-4 (VLA-4), lymphocyte function-associated antigen-1 (LFA-1) and macrophage-1 antigen (MAC-1) expression in U937 cells was upregulated by IL-17A. Furthermore, IL-17A stimulation resulted in mRNA and protein expression of scavenger receptor (LOX-1) in phorbol 12-myristate 13-acetate (PMA)-activated U937 cells. Oil Red O also demonstrated that IL-17A enhanced foam cell formation by PMA-activated U937 cells induced by oxidized low-density lipoprotein (ox-LDL), and this enhancement of ox-LDL-induced foam cell formation in IL-17A-treated U937 cells was downregulated by transfection of LOX-1 siRNA. These results indicated that IL-17A induced the expression of adhesion molecules, promoted the adherence of monocytes to vascular endothelial cells. IL-17A also stimulated ox-LDL-induced foam cell formation via upregulation of LOX-1 in activated macrophages. IL-17A may be responsible for the pathogenesis of atherosclerosis by inducing the adhesion of leukocytes to vascular endothelium and foam cell formation. [ABSTRACT FROM AUTHOR]

Seroepidemiological studies have highlighted a positive relation between CagA-positive Helicobacter pylori (H. pylori), atherosclerosis and related clinic events. However, this link has not been well validated. The present study was designed to explore the role of H. pylori PMSS1 (a CagA-positive strain that can translocate CagA into host cells) and exosomal CagA in the progression of atherosclerosis. To evaluate whether H. pylori accelerates or even induces atherosclerosis, H. pylori -infected C57/BL6 mice and ApoE−/− mice were maintained under different dietary conditions. To identify the role of H. pylori -infected gastric epithelial cells-derived exosomes (Hp -GES-EVs) and exosomal CagA in atherosclerosis, ApoE−/− mice were given intravenous or intraperitoneal injections of saline, GES-EVs, Hp -GES-EVs, and recombinant CagA protein (rCagA). CagA-positive H. pylori PMSS1 infection does not induce but promotes macrophage-derived foam cell formation and augments atherosclerotic plaque growth and instability in two animal models. Meanwhile, circulating Hp -GES-EVs are taken up in aortic plaque, and CagA is secreted in Hp -GES-EVs. Furthermore, the CagA-containing EVs and rCagA exacerbates macrophage-derived foam cell formation and lesion development in vitro and in vivo , recapitulating the pro-atherogenic effects of CagA-positive H. pylori. Mechanistically, CagA suppresses the transcription of cholesterol efflux transporters by downregulating the expression of transcriptional factors PPARγ and LXRα and thus enhances foam cell formation. These results may provide new insights into the role of exosomal CagA in the pathogenesis of CagA-positive H. pylori infection-related atherosclerosis. It is suggested that preventing and eradicating CagA-positive H. pylori infection could reduce the incidence of atherosclerosis and related events. • CagA-positive H. pylori infection does not initiate the development of atherosclerosis but can accelerate its progression by promoting macrophage-derived foam cell formation. • CagA-positive H. pylori infection promotes atherogenesis by releasing CagA-containing EVs and exosomal CagA mediates this effect. • CagA suppresses the transcription of cholesterol efflux transporters by downregulating the expression of transcriptional factors PPARγ and LXRα and thus enhances foam cell formation. [ABSTRACT FROM AUTHOR]