Yue Gu

Pulmonary hypertension is a severe vascular disease characterised by pulmonary arterial remodeling and right heart failure. However, the role of the glycolytic protein P4HA1 in its pathogenesis remains elusive. RNA-seq analysis was performed on hypoxic human pulmonary artery endothelial cells, and P4HA1 expression was validated in PH animal models. In vitro, P4HA1 knockdown was used to assess its effects on glycolysis, proliferation, and migration. Mechanistic studies included HIF1α transcriptional regulation and P4HA1-mediated HIF1α stabilization via reduced ubiquitination. AAV9-shP4HA1 was employed to assess P4HA1’s function in pulmonary vascular remodeling in vivo. P4HA1 was significantly upregulated in human pulmonary arterial endothelial cells (HPAECs) exposed to hypoxia. Mechanistic investigations revealed that HIF1α transcriptionally activates P4HA1 expression, while P4HA1 reciprocally stabilizes HIF1α by attenuating its ubiquitin-mediated degradation, thereby establishing a self-amplifying signaling loop. Functional studies demonstrated that P4HA1 knockdown effectively suppressed hypoxia-induced glycolytic reprogramming, inhibited cellular proliferation, and impaired migratory capacity in HPAECs. Consistent with these in vitro findings, endothelial-specific P4HA1 knockdown in vivo significantly attenuated hypoxia-induced pulmonary vascular remodeling and improved right ventricular function in a mouse PH model. Our studty identify P4HA1 as a novel contributor to PH pathogenesis and suggest that targeting the P4HA1/HIF1α axis may offer a promising therapeutic approach for pulmonary hypertension.

BACKGROUND: Vascular smooth muscle cell (VSMC) phenotypic modulation is responsible for the pathogenesis of hyper-muscularized arterial diseases. Recent studies have highlighted the critical role of epigenetic regulation in VSMC fate. However, the mechanisms underlying the precise regulation of the epigenetic machinery in VSMC remain unclear. METHODS: Using mouse aortic smooth muscle cells, carotid artery injury mouse model, and human atherosclerosis data sets, we identified GRK2 (G-protein–coupled receptor kinase 2) as a novel epigenetic regulator governing VSMC fate. RESULTS: GRK2 expression was found to be elevated in dedifferentiated VSMCs. Pharmacological or genetic silencing of GRK2 inhibited VSMC phenotypic switching. Mechanistic investigations demonstrated that GRK2 modulated VSMC phenotype via DNMT1 (DNA methyltransferase 1)-mediated DNA methylation. GRK2 phosphorylated DNMT1, stabilizing it by modulating its ubiquitination. Hypermethylated VSMC exhibited reduced expression of contractile-associated proteins. Inhibition of DNMT1 abolished the effects of GRK2 overexpression on VSMC phenotype, indicating a DNMT1-mediated mechanism. CONCLUSIONS: Our findings revealed that the GRK2-DNMT1 signaling axis is a critical regulator in VSMC phenotypic switching and present a potential therapeutic target for vascular remodeling.

Background: Vascular calcification (VC) is a common pathologic state that often accompanies calcium-phosphorus metabolism disorder and chronic kidney diseases (CKDs) in aging individuals. Vascular smooth muscle cell (VSMC) has been widely acknowledged as one of the main cell types that involved in this process. Niacin, a lipid-lowering reagent, has been demonstrated to be beneficial in atherosclerotic disease. The correlation and mechanism between niacin and vascular calcification have not been reported so far. Methods and results: RCS analysis based on large clinical databases indicated that diet that riches in niacin can protect against abdominal artery calcification (AAC). Our data showed that niacin treatment remarkably reduced VSMC osteogenic differentiation and senescence-associated markers under osteogenic stimulation. Moreover, niacin treatment alleviated CKD and vitamin D ₃ -induced vascular calcification in C57BL/6J mice. Mechanistically, we for the first time demonstrated that niacin supplementation inhibited vascular calcification via maintaining both Sirtuin 1 (SIRT1) and Sirtuin 6 (SIRT6) levels. Moreover, we verified that niacin activated SIRT1 and SIRT6 promoted VSMC autophagy flux. Conclusions: These findings may help to develop novel therapeutic strategies in the treatment and prevention of vascular calcification.

BACKGROUND: In-stent restenosis is characterized by a significant reduction in lumen diameter within the stented segment, primarily attributed to excessive proliferation of vascular smooth muscle cells (VSMCs) and neointimal hyperplasia. PFN1 (profilin-1), an actin-sequestering protein extensively studied in amyotrophic lateral sclerosis, remains less explored in neointimal hyperplasia. METHODS: Utilizing single-cell RNA sequencing alongside data from in-stent restenosis patients and various experimental in-stent restenosis models (swine, rats, and mice), we investigated the role of PFN1 in promoting VSMC phenotype switching and neointimal hyperplasia. RESULTS: Single-cell RNA sequencing of stenotic rat carotid arteries revealed a critical role for PFN1 in neointimal hyperplasia, a finding corroborated in stented swine coronary arteries, in-stent restenosis patients, PFN1 ^SMC-IKO (SMC-specific PFN1 knockout) mice, and PFN1 overexpressed mice. PFN1 deletion was shown to suppress VSMC phenotype switching and neointimal hyperplasia in PFN1 ^SMC-IKO mice subjected to a wire-injured model. To elucidate the observed discordance in PFN1 mRNA and protein levels, we identified that METTL3 (N ⁶ -methyladenosine methyltransferase) and YTHDF3 (YTH N6-methyladenosine RNA binding protein F3; N ⁶ -methyladenosine–specific reader) enhance PFN1 translation efficiency in an N ⁶ -methyladenosine–dependent manner, confirmed through experiments involving METTL3 knockout and YTHDF3 knockout mice. Furthermore, PFN1 was mechanistically found to interact with the phosphorylation of ANXA2 (annexin A2) by recruiting Src (SRC proto-oncogene, nonreceptor tyrosine kinase), promoting the phosphorylation of STAT3 (signal transducer and activator of transcription 3), a typical transcription factor known to induce VSMC phenotype switching. CONCLUSIONS: This study unveils the significance of PFN1 N ⁶ -methyladenosine modification in VSMCs, demonstrating its role in promoting phenotype switching and neointimal hyperplasia through the activation of the p-ANXA2 (phospho-ANXA2)/STAT3 pathway.

Artificial sweeteners are generally used and recommended to alternate added sugar for health promotion. However, the health effects of artificial sweeteners remain unclear. In this study, we included 6371 participants from the National Health and Nutrition Examination Survey with artificial sweetener intake records. Logistic regression and Cox regression were applied to explore the associations between artificial sweeteners and risks of cardiometabolic disorders and mortality. Mendelian randomisation was performed to verify the causal associations. We observed that participants with higher consumption of artificial sweeteners were more likely to be female and older and have above medium socio-economic status. After multivariable adjustment, frequent consumers presented the OR (95 % CI) for hypertension (1·52 (1·29, 1·80)), hypercholesterolaemia (1·28 (1·10, 1·50)), diabetes (3·74 (3·06, 4·57)), obesity (1·52 (1·29, 1·80)), congestive heart failure (1·89 (1·35, 2·62)) and heart attack (1·51 (1·10, 2·04)). Mendelian randomisation confirmed the increased risks of hypertension and type 2 diabetes. Moreover, an increased risk of diabetic mortality was identified in participants who had artificial sweeteners ≥ 1 daily (HR = 2·62 (1·46, 4·69), P = 0·001). Higher consumption of artificial sweeteners is associated with increased risks of cardiometabolic disorders and diabetic mortality. These results suggest that using artificial sweeteners as sugar substitutes may not be beneficial.

Doxorubicin (Dox) is a powerful antineoplastic agent, but its usage is limited by the severe cardiotoxicity referred to as Dox-induced cardiomyopathy (DIC). However, the molecular mechanism underlying this cardiotoxicity is yet to be fully elucidated. Here, our current study sought to determine the role of ubiquitin-specific protease 36 (USP36), a nucleolar deubiquitinating enzyme (DUB), in the progress of DIC and its mechanism. We identified an increased expression of USP36 both in neonatal rat cardiomyocytes and H9C2 cells exposed to Dox, and USP36 silencing significantly ameliorated Dox-induced oxidative stress injury and apoptosis in vitro. Mechanistically, USP36 upregulation was observed to positively correlate with PARP1 expression, and its knockdown resulted in reduction of PARP1 levels. Further investigation showed that USP36 could bind to and mediate the deubiquitination of PARP1 and increase its protein stability in cardiomyocytes upon Dox exposure. Moreover, overexpression of wild-type (WT) USP36 plasmid, but not its catalytic-inactive mutant (C131A), stabilizes PARP1 in HEK293T cells. Herein, we also established DIC model in mice and observed a significant upregulation of USP36 in the heart. Cardiac knockdown of USP36 in mice by a type 9 recombinant adeno-associated virus (rAAV9)-shUSP36 significantly preserved cardiac function after Dox treatment and protected against Dox-induced in terms of structural changes within the myocardium. Collectively, these findings indicate that Dox promotes DIC progression by activating USP36-mediated PARP1 deubiquitination. This novel USP36/PARP1 axis may play an important regulatory mechanism in the pathogenesis of DIC.