Long Noncoding RNA Gpr137b-ps Promotes Advanced Atherosclerosis via the Regulation of Autophagy in Macrophages.

BACKGROUND: Current therapies cannot completely reverse advanced atherosclerosis. High levels of amino acids, induced by Western diet, stimulate mTORC1 (mammalian target of rapamycin complex 1)-autophagy defects in macrophages, accelerating atherosclerotic plaque progression. In addition, autophagy-lysosomal dysfunction contributes to plaque necrotic core enlargement and lipid accumulation. Therefore, it is essential to investigate the novel mechanism and molecules to reverse amino acid-mTORC1-autophagy signaling dysfunction in macrophages of patients with advanced atherosclerosis. METHODS: We observed that Gpr137b-ps (G-protein-coupled receptor 137B, pseudogene) was upregulated in advanced atherosclerotic plaques. The effect of Gpr137b-ps on the progression of atherosclerosis was studied by generating advanced plaques in ApoE-/- mice with cardiac-specific knockout of Gpr137b-ps. Bone marrow-derived macrophages and mouse mononuclear macrophage cell line RAW264.7 cells were subjected to starvation or amino acid stimulation to study amino acid-mTORC1-autophagy signaling. Using both gain- and loss-of-function approaches, we explored the mechanism of Gpr137b-ps-regulated autophagy. RESULTS: Our results demonstrated that Gpr137b-ps deficiency led to enhanced autophagy in macrophages and reduced atherosclerotic lesions, characterized by fewer necrotic cores and less lipid accumulation. Knockdown of Gpr137b-ps increased autophagy and prevented amino acid-induced mTORC1 signaling activation. As the downstream binding protein of Gpr137b-ps, HSC70 (heat shock cognate 70) rescued the impaired autophagy induced by Gpr137b-ps. Furthermore, Gpr137b-ps interfered with the HSC70 binding to G3BP (Ras GTPase-activating protein-binding protein), which tethers the TSC (tuberous sclerosis complex) complex to lysosomes and suppresses mTORC1 signaling. In addition to verifying that the NTF2 (nuclear transport factor 2) domain of G3BP binds to HSC70 by in vitro protein synthesis, we further demonstrated that HSC70 binds to the NTF2 domain of G3BP through its W90-F92 motif by using computational modeling. CONCLUSIONS: These findings reveal that Gpr137b-ps plays an essential role in the regulation of macrophage autophagy, which is crucial for the progression of advanced atherosclerosis. Gpr137b-ps impairs the interaction of HSC70 with G3BP to regulate amino acid-mTORC1-autophagy signaling, and these results provide a new potential therapeutic direction for the treatment of advanced atherosclerosis.

Anti-atherosclerotic effects of geraniin through the gut microbiota-dependent trimethylamine N-oxide (TMAO) pathway in mice.

BACKGROUND: Cardiovascular disease is a leading cause of death, which signifies the urgent need for effective anti-atherosclerotic strategies. Gut microbiota-dependent trimethylamine-N-oxide (TMAO) is associated with atherosclerosis, and geraniin, a natural polyphenol with various biological activities, might play key role in this process. PURPOSE: We aimed to investigate the pharmacological activity of geraniin in atherosclerosis through remodeling the gut microbiota. METHODS: C57BL/6J ApoE-/- mice were administrated geraniin for 12 weeks. The colon contents were analyzed via 16S rRNA sequencing. Pathological staining was performed to evaluate the atherosclerotic characteristics. Cytokine assays detected the levels of plasma inflammatory cytokines. RAW264.7 cells were cultured in vitro and treated with TMAO. Tandem Mass Tag quantitative proteomics analysis and western blot were performed to investigate the effect of TMAO in macrophages. RESULTS: The plasma TMAO level in mice significantly decreased after geraniin intervention. The predominant intestinal microflora from geraniin-treated mice were Bacteroides (65.3%) and Firmicutes (30.6%). Pathological staining demonstrated that administration of geraniin attenuated atherosclerotic characteristics. After geraniin treatment, plasma levels of IL-1ß, IL-6, and TNF-α in mice were significantly reduced, and IL-10 levels were significantly increased. Proteomics analysis demonstrated the number of differentially expressed proteins after TMAO administration. In vitro study suggested that the atherogenic effect of TMAO could be attributed to changes in CD36, transmembrane protein 106a, apolipoprotein C1, macrophage scavenger receptor types I and II, and alpha-2-macroglobulin. CONCLUSION: Geraniin might be an effective prospective drug against cardiovascular diseases, and the gut microbiota is a potential target to reduce the risk of atherosclerotic disease.

Macrophage-Enriched lncRNA RAPIA: A Novel Therapeutic Target for Atherosclerosis.

OBJECTIVE: Despite the current antiatherosclerotic and antithrombotic therapies, the incidence of advanced atherosclerosis-associated clinical events remains high. Whether long noncoding RNAs (lncRNAs) affect the progression of atherosclerosis and whether they are potential targets for the treatment of advanced atherosclerosis are poorly understood. Approach and Results: The progression of atherosclerotic lesions was accompanied by dynamic alterations in lncRNA expression, as revealed by RNA sequencing and quantitative polymerase chain reaction. Among the dynamically changing lncRNAs, we identified a novel lncRNA, lncRNA Associated with the Progression and Intervention of Atherosclerosis (RAPIA), that was highly expressed in advanced atherosclerotic lesions and in macrophages. Inhibition of RAPIA in vivo not only repressed the progression of atherosclerosis but also exerted atheroprotective effects similar to those of atorvastatin on advanced atherosclerotic plaques that had already formed. In vitro assays demonstrated that RAPIA promoted proliferation and reduced apoptosis of macrophages. A molecular sponge interaction between RAPIA and microRNA-183-5p was demonstrated by dual-luciferase reporter and RNA immunoprecipitation assays. Rescue assays indicated that RAPIA functioned at least in part by targeting the microRNA-183-5p/ITGB1 (integrin ß1) pathway in macrophages. In addition, the transcription factor FoxO1 (forkhead box O1) could bind to the RAPIA promoter region and facilitate the expression of RAPIA. CONCLUSIONS: The progression of atherosclerotic lesions was accompanied by dynamic changes in the expression of lncRNAs. Inhibition of the pivotal lncRNA RAPIA may be a novel preventive and therapeutic strategy for advanced atherosclerosis, especially in patients resistant or intolerant to statins.