Application of New Lineage Tracing Techniques in Cardiovascular Development and Physiology.

Cardiovascular disease has been the leading cause of mortality and morbidity worldwide in the past 3 decades. Multiple cell lineages undergo dynamic alternations in gene expression, cell state determination, and cell fate conversion to contribute, adapt, and even modulate the pathophysiological processes during disease progression. There is an urgent need to understand the intricate cellular and molecular underpinnings of cardiovascular cell development in homeostasis and pathogenesis. Recent strides in lineage tracing methodologies have revolutionized our understanding of cardiovascular biology with the identification of new cellular origins, fates, plasticity, and heterogeneity within the cardiomyocyte, endothelial, and mesenchymal cell populations. In this review, we introduce the new technologies for lineage tracing of cardiovascular cells and summarize their applications in studying cardiovascular development, diseases, repair, and regeneration.

SARS-CoV-2 induced vascular endothelial dysfunction: direct or indirect effects?

Clinical evidence reveals that manifestations of endothelial dysfunction are widely observed in COVID-19 and long-COVID patients. However, whether these detrimental effects are caused by direct infection of the endothelium or are indirectly mediated by systemic inflammation has been a matter of debate. It has been well acknowledged that endothelial cells (ECs) of the cardiovascular system ubiquitously express the SARS-CoV-2 entry receptor angiotensin-converting enzyme 2 (ACE2), yet accumulating evidence suggests that it is more predominantly expressed by pericytes and vascular smooth muscle cells of the mammalian blood vessel. Besides, replicative infection of ECs by SARS-CoV-2 has yet to be demonstrated both in vitro and in vivo. In this study, we review latest research on endothelial ACE2 expression in different vascular beds, and the heterogeneity in various EC subsets with differential ACE2 expression in response to SARS-CoV-2. We also discuss ACE2-independent alternative mechanisms underlying endothelial activation in COVID-19, and the clinical manifestations of SARS-CoV-2-induced endothelial dysfunction. Altogether, understanding ACE2-dependent and ACE2-independent mechanisms driving SARS-CoV-2-induced vascular dysfunction would shed light on strategies of more effective therapies targeting cardiovascular complications associated with COVID-19.

Role of Argininosuccinate Synthase 1 -Dependent L-Arginine Biosynthesis in the Protective Effect of Endothelial Sirtuin 3 Against Atherosclerosis.

Atherosclerosis is initiated with endothelial cell (EC) dysfunction and vascular inflammation under hyperlipidemia. Sirtuin 3 (SIRT3) is a mitochondrial deacetylase. However, the specific role of endothelial SIRT3 during atherosclerosis remains poorly understood. The present study aims to study the role and mechanism of SIRT3 in EC function during atherosclerosis. Wild-type Sirt3f/f mice and endothelium-selective SIRT3 knockout Sirt3f/f; Cdh5Cre/+ (Sirt3EC-KO) mice are injected with adeno-associated virus (AAV) to overexpress PCSK9 and fed with high-cholesterol diet (HCD) for 12 weeks to induce atherosclerosis. Sirt3EC-KO mice exhibit increased atherosclerotic plaque formation, along with elevated macrophage infiltration, vascular inflammation, and reduced circulating L-arginine levels. In human ECs, SIRT3 inhibition resulted in heightened vascular inflammation, reduced nitric oxide (NO) production, increased reactive oxygen species (ROS), and diminished L-arginine levels. Silencing of SIRT3 results in hyperacetylation and deactivation of Argininosuccinate Synthase 1 (ASS1), a rate-limiting enzyme involved in L-arginine biosynthesis, and this effect is abolished in mutant ASS1. Furthermore, L-arginine supplementation attenuates enhanced plaque formation and vascular inflammation in Sirt3EC-KO mice. This study provides compelling evidence supporting the protective role of endothelial SIRT3 in atherosclerosis and also suggests a critical role of SIRT3-induced deacetylation of ASS1 by ECs for arginine synthesis.

Activation of Nrf2 inhibits atherosclerosis in ApoE-/- mice through suppressing endothelial cell inflammation and lipid peroxidation.

BACKGROUND: Nuclear erythroid 2-related factor 2 (Nrf2), a transcription factor, is critically involved in the regulation of oxidative stress and inflammation. However, the role of endothelial Nrf2 in atherogenesis has yet to be defined. In addition, how endothelial Nrf2 is activated and whether Nrf2 can be targeted for the prevention and treatment of atherosclerosis is not explored. METHODS: RNA-sequencing and single-cell RNA sequencing analysis of mouse atherosclerotic aortas were used to identify the differentially expressed genes. In vivo endothelial cell (EC)-specific activation of Nrf2 was achieved by injecting adeno-associated viruses into ApoE-/- mice, while EC-specific knockdown of Nrf2 was generated in Cdh5CreCas9floxed-stopApoE-/- mice. RESULTS: Endothelial inflammation appeared as early as on day 3 after feeding of a high cholesterol diet (HCD) in ApoE-/- mice, as reflected by mRNA levels, immunostaining and global mRNA profiling, while the immunosignal of the end-product of lipid peroxidation (LPO), 4-hydroxynonenal (4-HNE), started to increase on day 10. TNF-α, 4-HNE, and erastin (LPO inducer), activated Nrf2 signaling in human ECs by increasing the mRNA and protein expression of Nrf2 target genes. Knockdown of endothelial Nrf2 resulted in augmented endothelial inflammation and LPO, and accelerated atherosclerosis in Cdh5CreCas9floxed-stopApoE-/- mice. By contrast, both EC-specific and pharmacological activation of Nrf2 inhibited endothelial inflammation, LPO, and atherogenesis. CONCLUSIONS: Upon HCD feeding in ApoE-/- mice, endothelial inflammation is an earliest event, followed by the appearance of LPO. EC-specific activation of Nrf2 inhibits atherosclerosis while EC-specific knockdown of Nrf2 results in the opposite effect. Pharmacological activators of endothelial Nrf2 may represent a novel therapeutic strategy for the treatment of atherosclerosis.