WTAP Promotes Atherosclerosis by Inducing Macrophage Pyroptosis and M1 Polarization through Upregulating NLRP3

Wang J., Ran Y., Li Z., Zhao T., Zhang F., Wang J., Liu Z., Chen X.

JOURNAL/nrgr/04.03/01300535-202503000-00032/figure1/v/2024-06-17T092413Z/r/image-tiff Salsolinol (1-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline, Sal) is a catechol isoquinoline that causes neurotoxicity and shares structural similarity with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, an environmental toxin that causes Parkinson’s disease. However, the mechanism by which Sal mediates dopaminergic neuronal death remains unclear. In this study, we found that Sal significantly enhanced the global level of N6-methyladenosine (m6A) RNA methylation in PC12 cells, mainly by inducing the downregulation of the expression of m6A demethylases fat mass and obesity-associated protein (FTO) and alkB homolog 5 (ALKBH5). RNA sequencing analysis showed that Sal downregulated the Hippo signaling pathway. The m6A reader YTH domain-containing family protein 2 (YTHDF2) promoted the degradation of m6A-containing Yes-associated protein 1 (YAP1) mRNA, which is a downstream key effector in the Hippo signaling pathway. Additionally, downregulation of YAP1 promoted autophagy, indicating that the mutual regulation between YAP1 and autophagy can lead to neurotoxicity. These findings reveal the role of Sal on m6A RNA methylation and suggest that Sal may act as an RNA methylation inducer mediating dopaminergic neuronal death through YAP1 and autophagy. Our results provide greater insights into the neurotoxic effects of catechol isoquinolines compared with other studies and may be a reference for assessing the involvement of RNA methylation in the pathogenesis of Parkinson’s disease.

Zhu J., Pan S., Chai H., Zhao P., Feng Y., Cheng Z., Zhang S., Wang W.

AbstractSingle‐cell mass spectrometry (MS) is significant in biochemical analysis and holds great potential in biomedical applications. Efficient sample preparation like sorting (i.e., separating target cells from the mixed population) and desalting (i.e., moving the cells off non‐volatile salt solution) is urgently required in single‐cell MS. However, traditional sample preparation methods suffer from complicated operation with various apparatus, or insufficient performance. Herein, a one‐step sample preparation strategy by leveraging label‐free impedance flow cytometry (IFC) based microfluidics is proposed. Specifically, the IFC framework to characterize and sort single‐cells is adopted. Simultaneously with sorting, the target cell is transferred from the local high‐salinity buffer to the MS‐compatible solution. In this way, one‐step sorting and desalting are achieved and the collected cells can be directly fed for MS analysis. A high sorting efficiency (>99%), cancer cell purity (≈87%), and desalting efficiency (>99%), and the whole workflow of impedance‐based separation and MS analysis of normal cells (MCF‐10A) and cancer cells (MDA‐MB‐468) are verified. As a standalone sample preparation module, the microfluidic chip is compatible with a variety of MS analysis methods, and envisioned to provide a new paradigm in efficient MS sample preparation, and further in multi‐modal (i.e., electrical and metabolic) characterization of single‐cells.

Lu W., Yang X., Zhong W., Chen G., Guo X., Ye Q., Xu Y., Qi Z., Ye Y., Zhang J., Wang Y., Wang X., Wang S., Zhao Q., Zeng W., et. al.

Tian S., Chen X., Wu W., Lin H., Qing X., Liu S., Wang B., Xiao Y., Shao Z., Peng Y.

AbstractLower back pain (LBP), which is a primary cause of disability, is largely attributed to intervertebral disc degeneration (IDD). Macrophages (MΦs) in degenerated intervertebral discs (IVDs) form a chronic inflammatory microenvironment, but how MΦs are recruited to degenerative segments and transform into a proinflammatory phenotype remains unclear. We evaluated chemokine expression in degenerated nucleus pulposus cells (NPCs) to clarify the role of NPCs in the establishment of an inflammatory microenvironment in IDD and explored the mechanisms. We found that the production of C-C motif chemokine ligand 2 (CCL2) and C-C motif chemokine ligand 7 (CCL7) was significantly increased in NPCs under inflammatory conditions, and blocking CCL2/7 and their receptor, C-C chemokine receptor type 2(CCR2), inhibited the inductive effects of NPCs on MΦ infiltration and proinflammatory polarization. Moreover, activation of the integrated stress response (ISR) was obvious in IDD, and ISR inhibition reduced the production of CCL2/7 in NPCs. Further investigation revealed that activating Transcription Factor 3 (ATF3) responded to ISR activation, and ChIP-qPCR verified the DNA-binding activity of ATF3 on CCL2/7 promoters. In addition, we found that Toll-like receptor 4 (TLR4) inhibition modulated ISR activation, and TLR4 regulated the accumulation of mitochondrial reactive oxygen species (mtROS) and double-stranded RNA (dsRNA). Downregulating the level of mtROS reduced the amount of dsRNA and ISR activation. Deactivating the ISR or blocking CCL2/7 release alleviated inflammation and the progression of IDD in vivo. Moreover, MΦ infiltration and IDD were inhibited in CCR2-knockout mice. In conclusion, this study highlights the critical role of TLR4/mtROS/dsRNA axis-mediated ISR activation in the production of CCL2/7 and the progression of IDD, which provides promising therapeutic strategies for discogenic LBP.

Wu J., Pan J., Zhou W., Ji G., Dang Y.

RNA methylation modifications, as a widespread type of modification in eukaryotic cells, especially N6-methyladenosine (m6A), are associated with many activities in organisms, including macrophage polarization and progression of non-alcoholic steatohepatitis (NASH). Macrophages in the liver are of diverse origin and complex phenotype, exhibiting different functions in development of NASH. In the review, we discuss the functions of m6A and m6A-related enzymes in macrophage polarization. Furthermore, we retrospect the role of macrophage polarization in NASH. Finally, we discuss the prospects of m6A in macrophages and NASH, and provide guidance for the treatment of NASH.

Zhao W., Xu Y., Zhu J., Zhang C., Zhou W., Wang S.

AbstractRNA N6-methyladenosine (m6A) regulators play essential roles in diverse biological processes, including immune responses. Mounting evidence suggests that their dysregulation is intricately linked to numerous diseases. However, the role of m6A-associated genes in carotid atherosclerosis and their relationship with aging and immune cells remain unclear. Analyze the expression profiles of m6A-related genes in carotid atherosclerosis-related datasets. Based on the expression patterns of m6A-related genes, perform consistent clustering analysis of carotid atherosclerosis samples and investigate associated immune cell infiltration patterns and aging characteristics. Develop an m6A prediction model specific to carotid atherosclerosis and analyze the relationships between immune cells infiltration and aging features. The m6A methylation modification level exhibited a substantial decrease in early-stage carotid atherosclerosis samples compared to late-stage carotid atherosclerosis samples. Subsequently, two distinct m6A subtypes were defined through consensus clustering analysis, with the lower m6A modification level group showing associations with heightened immune cell infiltration and increased expression of aging-related genes. A model composed of five m6A-related genes was formulated, and the results indicated that this model possesses effective predictive and therapeutic capabilities for carotid atherosclerosis. Furthermore, the downregulation of YTHDC1 expression resulted in elevated expression of inflammatory factors and a decrease in the expression of the aging-related gene RGN. Single-cell data analysis suggests that the reduced expression of YTHDC1 may decrease the degradation of inflammation-related factors in macrophages, leading to a highly inflammatory state in the carotid artery wall. Furthermore, the sustained release of inflammatory factors may increase the expression of the aging-related gene RGN in vascular smooth muscle cells, further exacerbating the progression of atherosclerosis. A reduced level of m6A methylation modification could enhance inflammation and expedite cellular aging, thereby contributing to the development of carotid atherosclerosis.

Wang P., Xie D., Xiao T., Cheng C., Wang D., Sun J., Wu M., Yang Y., Zhang A., Liu Q.

As epigenetic modifications, lactylation and N6-methyladenosine (m6A) have attracted wide attention. Arsenite is an environmental pollutant that has been proven to induce idiopathic pulmonary fibrosis (IPF). However, the molecular mechanisms of lactylation and m6A methylation are unclear in arsenite-related IPF (As-IPF). In view of the limited understanding of molecular mechanism of m6A and lactylation in As-IPF, MeRIP-seq, RNA-seq and ChIP-seq were analyzed to verify the target gene regulated by m6A and H3K18 lactylation (H3K18la). We found that, for As-IPF, the global levels of m6A, levels of YTHDF1 and m6A-modified neuronal protein 3.1 (NREP) were elevated in alveolar epithelial cells (AECs). The secretion levels of TGF-β1 were increased via YTHDF1/m6A/NREP, which promoted the fibroblast-to-myofibroblast transition (FMT). Further, extracellular lactate from myofibroblasts elevated levels of the global lactylation (Kla) and H3K18la via the lactate monocarboxylate transporter 1 (MCT1), and, in AECs, H3K18la facilitated the transcription of Ythdf1. This report highlights the role of crosstalk between AECs and myofibroblasts via lactylation and m6A and the significance of H3K18la regulation of YTHDF1 in the progression of As-IPF, which may be useful for finding effective therapeutic targets.

Li B., Wang W., Zhao L., Li M., Yan D., Li X., Zhang J., Gao Q., Feng Y., Zheng J., Shu B., Yan Y., Wang J., Wang H., He L., et. al.

AbstractThe recent prevalence of monkeypox has led to the declaration of a public health emergency of international concern. Monkeypox lesions are typically ulcers or pustules (containing high titers of replication‐competent virus) in the skin and mucous membranes, which allows monkeypox virus to transmit predominantly through intimate contact. Currently, effective clinical treatments for monkeypox are lacking, and strategies for blocking virus transmission are fraught with drawbacks. Herein, we construct a biomimetic nanotemplate (termed TBD@M NPs) with macrophage membranes as the coat and polymeric nanoparticles loading a versatile aggregation‐induced emission (AIE)‐featured photothermal molecule TPE‐BT‐DPTQ as the core. In a surrogate mouse model of monkeypox (vaccinia virus‐infected tail scarification model), intravenously injected TBD@M NPs show precise tracking and near‐infrared region II (NIR‐II) fluorescence imaging (FLI) of the lesions. Upon 808 nm laser irradiation, the virus is eliminated by the photothermal effect and the infected wound heals rapidly. More importantly, the inoculation of treated lesion tissue suspensions does not trigger tail infection or inflammatory activation in healthy mice, indicating successful blockage of virus transmission. This study demonstrates for the first time monkeypox theranostics using nanomedicine, and may bring a new insight into the development of a viable strategy for monkeypox management in clinical trials.This article is protected by copyright. All rights reserved

Sun T., Lv J., Zhao X., Li W., Zhang Z., Nie L.

We present a rapid and high-resolution photoacoustic imaging method for evaluating the liver function reserve (LFR). To validate its accuracy, we establish alcoholic liver disease (ALD) models and employ dual-wavelength spectral unmixing to assess oxygen metabolism. An empirical mathematical model fits the photoacoustic signals, obtaining liver metabolism curve and LFR parameters. Liver oxygen metabolism significantly drops in ALD with the emergence of abnormal hepatic lobular structure. ICG half-life remarkably extends from 241 to 568 s in ALD. A significant decline in LFR occurs in terminal region compared to central region, indicated by a 106.9 s delay in ICG half-life, likely due to hepatic artery and vein damage causing hypoxia and inadequate nutrition. Reduced glutathione repairs LFR with a 43% improvement by reducing alcohol-induced oxidative damage. Scalable photoacoustic imaging shows immense potential for assessing LFR in alcoholic-related diseases, providing assistance to early detection and management of liver disease.

Hou P., Fang J., Liu Z., Shi Y., Agostini M., Bernassola F., Bove P., Candi E., Rovella V., Sica G., Sun Q., Wang Y., Scimeca M., Federici M., Mauriello A., et. al.

AbstractAtherosclerosis is a chronic inflammatory disease characterized by the accumulation of fatty deposits in the inner walls of vessels. These plaques restrict blood flow and lead to complications such as heart attack or stroke. The development of atherosclerosis is influenced by a variety of factors, including age, genetics, lifestyle, and underlying health conditions such as high blood pressure or diabetes. Atherosclerotic plaques in stable form are characterized by slow growth, which leads to luminal stenosis, with low embolic potential or in unstable form, which contributes to high risk for thrombotic and embolic complications with rapid clinical onset. In this complex scenario of atherosclerosis, macrophages participate in the whole process, including the initiation, growth and eventually rupture and wound healing stages of artery plaque formation. Macrophages in plaques exhibit high heterogeneity and plasticity, which affect the evolving plaque microenvironment, e.g., leading to excessive lipid accumulation, cytokine hyperactivation, hypoxia, apoptosis and necroptosis. The metabolic and functional transitions of plaque macrophages in response to plaque microenvironmental factors not only influence ongoing and imminent inflammatory responses within the lesions but also directly dictate atherosclerotic progression or regression. In this review, we discuss the origin of macrophages within plaques, their phenotypic diversity, metabolic shifts, and fate and the roles they play in the dynamic progression of atherosclerosis. It also describes how macrophages interact with other plaque cells, particularly T cells. Ultimately, targeting pathways involved in macrophage polarization may lead to innovative and promising approaches for precision medicine. Further insights into the landscape and biological features of macrophages within atherosclerotic plaques may offer valuable information for optimizing future clinical treatment for atherosclerosis by targeting macrophages.

Li J., Zou C., Zhang Z., Xue F.

Cerebrovascular diseases have extreme high mortality and disability rate worldwide, and endothelial cells injury-induced atherosclerosis acts as the main cause of cerebrovascular disease. Ferroptosis is a novel type of programmed cell death depending on iron-lipid peroxidation. Recent studies have revealed that ferroptosis might promote the progression of atherosclerosis (AS). Here, this research aimed to investigate the function and its profound mechanism on vascular endothelial cells in atherosclerosis. Research results revealed that YTHDF2 expression up-regulated in ox-LDL treated human umbilical vein endothelial cells (HUVECs). Gain/loss functional assays indicated that YTHDF2 overexpression inhibited HUVECs’ proliferation and accelerated the ferroptosis in ox-LDL-administered HUVECs. Meanwhile, YTHDF2 silencing promoted cell proliferation and reduced the ferroptosis in ox-LDL-administered HUVECs. Mechanistically, in silico analysis suggested that there were potential m6A-modified sites on SLC7A11 mRNA, and YTHDF2 could bind with SLC7A11 mRNA via m6A-dependent manner. YTHDF2 promoted the degradation of SLC7A11 mRNA, thereby reducing its mRNA stability. Taken together, these findings suggest that YTHDF2 accelerates endothelial cells ferroptosis in cerebrovascular atherosclerosis, helping us enhance our comprehension on cerebrovascular disease pathological physiology.

Liu X., Luo P., Zhang W., Zhang S., Yang S., Hong F.

Pyroptosis is a pro-inflammatory type of regulated cell death (RCD) characterized by gasdermin protein-mediated membrane pore formation, cell swelling, and rapid lysis. Recent studies have suggested that pyroptosis is closely related to atherosclerosis (AS). Previous studies reported that pyroptosis involving endothelial cells (ECs), macrophages, and smooth muscle cells (SMCs) plays an important role in the formation and development of AS. Pyroptosis not only causes local inflammation but also amplifies the inflammatory response and it aggravates plaque instability, leading to plaque rupture and thrombosis, eventually resulting in acute cardiovascular events. In this review, we clarified some novel pathways and mechanics and presented some potential drugs.

Yan H., Huang W., Rao J., Yan D., Yuan J.

Neuronal death following ischemia is the primary cause of death and disability in patients with ischemic stroke. N6-methyladenosine (m6A) modification plays essential role in various physiological and pathological conditions, but its role and mechanism in ischemic neuronal death remain unclear. In the present study, neuronal pyroptosis was an important event in brain injury caused by ischemic stroke, and the upregulation of long non-coding RNA (lncRNA) maternally expressed gene 3 (MEG3) following cerebral ischemia was a key factor in activating ischemic neuronal pyroptosis via NLRP3/caspase-1/GSDMD signaling. Moreover, we first demonstrated that the demethylase fat mass and obesity-associated protein (FTO), which was decreased following ischemia, regulated MEG3 expression in an m6A-dependent manner by affecting its stability, thereby activating neuronal pyroptosis via NLRP3/caspase-1/GSDMD signaling, and ultimately leading to ischemic brain damage. Therefore, the present study provides new insights for the mechanism of ischemic stroke, and suggests that FTO may be a potential therapeutic target for ischemic stroke.

Morrison A.M., Sullivan A.E., Aday A.W.

Atherosclerotic disease, including stroke and myocardial infarction, is the leading cause of morbidity and mortality worldwide. Atherosclerotic plaque formation occurs in the setting of excess oxidative and hemodynamic stress and is perpetuated by smoking, poor diet, dyslipidemia, hypertension, and diabetes. Plaque may rupture, resulting in acute thrombotic events. Smoking cessation, lifestyle modification, risk factor optimization, and antithrombotic therapies are the mainstays of atherosclerotic disease management and are the cornerstones to reduce morbidity and mortality in this high-risk patient population. Novel therapeutics are in development and will add to the growing armamentarium available to physicians who manage atherosclerotic disease.

Pan W., Zhang J., Zhang L., Zhang Y., Song Y., Han L., Tan M., Yin Y., Yang T., Jiang T., Li H.

AbstractCardiovascular diseases (CVDs) are the primary drivers of the growing public health epidemic and the leading cause of premature mortality and economic burden worldwide. With decades of research, CVDs have been proven to be associated with the dysregulation of the inflammatory response, with macrophages playing imperative roles in influencing the prognosis of CVDs. Autophagy is a conserved pathway that maintains cellular functions. Emerging evidence has revealed an intrinsic connection between autophagy and macrophage functions. This review focuses on the role and underlying mechanisms of autophagy‐mediated regulation of macrophage plasticity in polarization, inflammasome activation, cytokine secretion, metabolism, phagocytosis, and the number of macrophages. In addition, autophagy has been shown to connect macrophages and heart cells. It is attributed to specific substrate degradation or signalling pathway activation by autophagy‐related proteins. Referring to the latest reports, applications targeting macrophage autophagy have been discussed in CVDs, such as atherosclerosis, myocardial infarction, heart failure, and myocarditis. This review describes a novel approach for future CVD therapies.