Renyang Tong

Abstract Rupture of vulnerable plaques is the main trigger of acute cardio‐cerebral vascular events, but mechanisms responsible for transforming a stable atherosclerotic into a vulnerable plaque remain largely unknown. Melatonin, an indoleamine hormone secreted by the pineal gland, plays pleiotropic roles in the cardiovascular system; however, the effect of melatonin on vulnerable plaque rupture and its underlying mechanisms remains unknown. Here, we generated a rupture‐prone vulnerable carotid plaque model induced by endogenous renovascular hypertension combined with low shear stress in hypercholesterolemic ApoE −/− mice. Melatonin (10 mg/kg/d by oral administration for 9 weeks) significantly prevented vulnerable plaque rupture, with lower incidence of intraplaque hemorrhage (42.9% vs. 9.5%, P = 0.014) and of spontaneous plaque rupture with intraluminal thrombus formation (38.1% vs. 9.5%, P = 0.029). Mechanistic studies indicated that melatonin ameliorated intraplaque inflammation by suppressing the differentiation of intraplaque macrophages toward the proinflammatory M1 phenotype, and circadian nuclear receptor retinoid acid receptor‐related orphan receptor‐α (RORα) mediated melatonin‐exerted vasoprotection against vulnerable plaque instability and intraplaque macrophage polarization. Further analysis in human monocyte‐derived macrophages confirmed the role of melatonin in regulating macrophage polarization by regulating the AMPKα‐STATs pathway in a RORα‐dependent manner. In summary, our data provided the first evidence that melatonin‐RORα axis acts as a novel endogenous protective signaling pathway in the vasculature, regulates intraplaque inflammation, and stabilizes rupture‐prone vulnerable plaques.

A comprehensive understanding of the evolution of the immune landscape in humans across the entire lifespan at single-cell transcriptional and protein levels, during development, maturation and senescence is currently lacking. We recruited a total of 220 healthy volunteers from the Shanghai Pudong Cohort (NCT05206643), spanning 13 age groups from 0 to over 90 years, and profiled their peripheral immune cells through single-cell RNA-sequencing coupled with single T cell and B cell receptor sequencing, high-throughput mass cytometry, bulk RNA-sequencing and flow cytometry validation experiments. We revealed that T cells were the most strongly affected by age and experienced the most intensive rewiring in cell–cell interactions during specific age. Different T cell subsets displayed different aging patterns in both transcriptomes and immune repertoires; examples included GNLY+CD8+ effector memory T cells, which exhibited the highest clonal expansion among all T cell subsets and displayed distinct functional signatures in children and the elderly; and CD8+ MAIT cells, which reached their peaks of relative abundance, clonal diversity and antibacterial capability in adolescents and then gradually tapered off. Interestingly, we identified and experimentally verified a previously unrecognized ‘cytotoxic’ B cell subset that was enriched in children. Finally, an immune age prediction model was developed based on lifecycle-wide single-cell data that can evaluate the immune status of healthy individuals and identify those with disturbed immune functions. Our work provides both valuable insights and resources for further understanding the aging of the immune system across the whole human lifespan. In this Resource, authors profile peripheral immune cells from a total of 220 healthy volunteers from birth to over 90 years. This revealed that T cells were most affected by aging with divergent aging patterns in different subsets and identified a population of cytotoxic B cells that were enriched in children.

Abstract Aims Adverse cardiovascular events have day/night patterns with peaks in the morning, potentially related to endogenous circadian clock control of platelet activation. Circadian nuclear receptor Rev-erbα is an essential and negative component of the circadian clock. To date, the expression profile and biological function of Rev-erbα in platelets have never been reported. Methods and results Here, we report the presence and functions of circadian nuclear receptor Rev-erbα in human and mouse platelets. Both human and mouse platelet Rev-erbα showed a circadian rhythm that positively correlated with platelet aggregation. Global Rev-erbα knockout and platelet-specific Rev-erbα knockout mice exhibited defective in haemostasis as assessed by prolonged tail-bleeding times. Rev-erbα deletion also reduced ferric chloride-induced carotid arterial occlusive thrombosis, prevented collagen/epinephrine-induced pulmonary thromboembolism, and protected against microvascular microthrombi obstruction and infarct expansion in an acute myocardial infarction model. In vitro thrombus formation assessed by CD41-labelled platelet fluorescence intensity was significantly reduced in Rev-erbα knockout mouse blood. Platelets from Rev-erbα knockout mice exhibited impaired agonist-induced aggregation responses, integrin αIIbβ3 activation, and α-granule release. Consistently, pharmacological inhibition of Rev-erbα by specific antagonists decreased platelet activation markers in both mouse and human platelets. Mechanistically, mass spectrometry and co-immunoprecipitation analyses revealed that Rev-erbα potentiated platelet activation via oligophrenin-1-mediated RhoA/ERM (ezrin/radixin/moesin) pathway. Conclusion We provided the first evidence that circadian protein Rev-erbα is functionally expressed in platelets and potentiates platelet activation and thrombus formation. Rev-erbα may serve as a novel therapeutic target for managing thrombosis-based cardiovascular disease. Key question Adverse cardiovascular events have day/night patterns with peaks in the morning, potentially related to endogenous circadian clock control of platelet activation. Whether circadian nuclear receptor Rev-erba is present in platelets and regulates platelet function remains unknown. Key finding We provide the first evidence that Rev-erba is functionally expressed in platelets and acts as a positive regulator of platelet activation/thrombus formation through the oligophrenin-1-mediated RhoA/ERM signalling pathway. Take home message Our observations highlight the importance of circadian clock machinery in platelet physiology and support the notion that Rev-erba may serve as a novel therapeutic target for managing thrombosis-based cardiovascular diseases.

Exercise-induced physiological hypertrophy provides protection against cardiovascular disease, whereas disease-induced pathological hypertrophy leads to heart failure. Emerging evidence suggests pleiotropic roles of melatonin in cardiac disease; however, the effects of melatonin on physiological vs pathological cardiac hypertrophy remain unknown. Using swimming-induced physiological hypertrophy and pressure overload-induced pathological hypertrophy models, we found that melatonin treatment significantly improved pathological hypertrophic responses accompanied by alleviated oxidative stress in myocardium but did not affect physiological cardiac hypertrophy and oxidative stress levels. As an important mediator of melatonin, the retinoid-related orphan nuclear receptor-α (RORα) was significantly decreased in human and murine pathological hypertrophic cardiomyocytes, but not in swimming-induced physiological hypertrophic murine hearts. In vivo and in vitro loss-of-function experiments indicated that RORα deficiency significantly aggravated pathological cardiac hypertrophy, and notably weakened the anti-hypertrophic effects of melatonin. Mechanistically, RORα mediated the cardioprotection of melatonin in pathological hypertrophy mainly by transactivation of manganese-dependent superoxide dismutase (MnSOD) via binding to the RORα response element located in the promoter region of the MnSOD gene. Furthermore, MnSOD overexpression reversed the pro-hypertrophic effects of RORα deficiency, while MnSOD silencing abolished the anti-hypertrophic effects of RORα overexpression in pathological cardiac hypertrophy. Collectively, our findings provide the first evidence that melatonin exerts an anti-hypertrophic effect on pathological but not physiological cardiac hypertrophy via alleviating oxidative stress through transactivation of the antioxidant enzyme MnSOD in a RORα-dependent manner.

Nonalcoholic fatty liver disease (NAFLD), characterized by hepatic steatosis (HS), insulin resistance (IR), and inflammation, poses a high risk of cardiometabolic disorders. Ubiquitin specific protease 4 (USP4), a deubiquitinating enzyme, is pivotally involved in regulating multiple inflammatory pathways; however, the role of USP4 in NAFLD is unknown. Here, we report that USP4 expression was dramatically down-regulated in livers from NAFLD patients and different NAFLD mouse models induced by high-fat diet (HFD) or genetic deficiency (ob/ob) as well as in palmitate-treated hepatocytes. Hepatocyte-specific USP4 depletion exacerbated HS, IR, and inflammatory response in HFD-induced NAFLD mice. Conversely, hepatic USP4 overexpression notably alleviated the pathological alterations in two different NAFLD models. Mechanistically, hepatocyte USP4 directly bound to and deubiquitinated transforming growth factor-β activated kinase 1 (TAK1), leading to a suppression of the activation of downstream nuclear factor kappa B (NF-κB) and c-Jun N-terminal kinase (JNK) cascades, which, in turn, reversed the disruption of insulin receptor substrate/protein kinase B/glycogen synthase kinase 3 beta (IRS-AKT-GSK3β) signaling. In addition, USP4-TAK1 interaction and subsequent TAK1 deubiquitination were required for amelioration of metabolic dysfunctions. Conclusion: Collectively, the present study provides evidence that USP4 functions as a pivotal suppressor in NAFLD and related metabolic disorders. (Hepatology 2018; 00:000-000).