Angiotensin-II (Ang-II) drives pathological vascular wall remodeling in hypertension and abdominal aortic aneurysm (AAA) through mechanisms that are not completely understood. Previous studies showed that the phosphatase activity of calcineurin (Cn) mediates Ang-II-induced AAA, but the cell type involved in the action of Cn in AAA formation remained unknown. Here, by employing newly created smooth muscle cell (SMC)-specific and endothelial cell (EC)-specific Cn-deficient mice (SM-Cn-/- and EC-Cn-/- mice), we show that Cn expressed in SMCs, but not ECs, was required for Ang-II-induced AAA. Unexpectedly, SMC Cn also played a structural role in the early onset and maintenance of Ang-II-induced hypertension, independently of its known phosphatase activity. Among the signaling pathways activated by Ang-II, Cn signaling is essential in SMCs, as nearly 90% of the genes regulated by Ang-II in the aorta required Cn expression in SMCs. Cn orchestrated, independently of its enzymatic activity, the induction by Ang-II of a transcriptional program closely related to SMC contractility and hypertension. Cn deletion in SMCs, but not its pharmacological inhibition, impaired the regulation of arterial contractility. Among the genes whose regulation by Ang-II required Cn expression but not its phosphatase activity, we discovered that Serpine1 was critical for Ang-II-induced hypertension. Indeed, pharmacological inhibition of PAI-1, the protein encoded by Serpine1, impaired SMCs contractility and readily regressed hypertension. Mechanistically, Serpine1 induction was mediated by Smad2 activation via the structural role of Cn. These findings uncover an unexpected role for Cn in vascular pathophysiology and highlight PAI-1 as a potential therapeutic target for hypertension.
Copyright: © 2025 Yunes-Leites et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Abstract Blood vessels provide a versatile and adaptable transport system, but recent work has established that endothelial cells, which form the innermost lining of the vascular network, are also a source of molecular signals controlling the behavior of other cell types in the surrounding tissue. Pericytes are another essential component of the vessel wall, but comparably little is known about their signaling interactions with other cell populations during organ growth and patterning. Here, we have used tissue-specific and inducible mouse genetics, high-resolution imaging, single-cell RNA sequencing and cell culture experiments to address the function of three pericyte-derived growth factors in the postnatal development of two model organs, namely lung and brain. We found that Pdgfrb-CreERT2-controlled inactivation of the gene for hepatocyte growth factor (HGF) causes no overt alterations in the postnatal brain but impairs alveologenesis in the lung due to defective interaction with AT2 epithelial cells. Likewise, expression of brain-derived neurotrophic factor (BDNF) by pericytes is not required in the postnatal brain but controls lung development through interactions with the receptor tyrosine kinase TrkB in the pulmonary endothelium. Conversely, pericyte expression of the TGFβ family growth factor Nodal is not required for lung morphogenesis but regulates blood vessel growth and barrier function in the postnatal brain, which we attribute to signaling interactions with endothelial cells, astrocytes and microglia. Taken together, our findings establish that pericytes are a critical source of angiocrine signals that control morphogenetic processes in an organ-specific fashion.

Atherosclerosis is a chronic inflammatory disease characterized by the accumulation of lipids and leukocytes within the arterial wall. By studying the aortic transcriptome of atherosclerosis-prone apolipoprotein E (ApoE-/-) mice, we aimed to identify novel players in the progression of atherosclerosis.
RNA-Seq analysis was performed on aortas from ApoE-/- and wild-type mice. AnxA8 expression was assessed in human and mice atherosclerotic tissue and healthy aorta. ApoE-/- mice lacking systemic AnxA8 (ApoE-/-AnxA8-/-) were generated to assess the effect of AnxA8 deficiency on atherosclerosis. Bone marrow transplantation (BMT) was also performed to generate ApoE-/- lacking AnxA8 specifically in bone marrow-derived cells. Endothelial-specific AnxA8 silencing in vivo was performed in ApoE-/- mice. The functional role of AnxA8 was analysed in cultured murine cells.
RNA-Seq unveiled AnxA8 as one of the most significantly upregulated genes in atherosclerotic aortas of ApoE-/- compared to wild-type mice. Moreover, AnxA8 was upregulated in human atherosclerotic plaques. Germline deletion of AnxA8 decreased the atherosclerotic burden, the size and volume of atherosclerotic plaques in the aortic root. Plaques of ApoE-/-AnxA8-/- were characterized by lower lipid and inflammatory content, smaller necrotic core, thicker fibrous cap and less apoptosis compared with those in ApoE-/-AnxA8+/+. BMT showed that hematopoietic AnxA8 deficiency had no effect on atherosclerotic progression. Oxidized low-density lipoprotein (ox-LDL) increased AnxA8 expression in murine aortic endothelial cells (MAECs). In vitro experiments revealed that AnxA8 deficiency in MAECs suppressed P/E-selectin and CD31 expression and secretion induced by ox-LDL with a concomitant reduction in platelet and leukocyte adhesion. Intravital microscopy confirmed the reduction in leukocyte and platelet adhesion in ApoE-/-AnxA8-/- mice. Finally, endothelial-specific silencing of AnxA8 decreased atherosclerosis progression.
Our findings demonstrate that AnxA8 promotes the progression of atherosclerosis by modulating endothelial-leukocyte interactions. Interventions capable of reducing AnxA8 expression in endothelial cells may delay atherosclerotic plaque progression.
This study shows that AnxA8 is upregulated in aorta of atheroprone mice and in human atherosclerotic plaques. Germline AnxA8 deficiency reduces platelet and leukocyte recruitment to activated endothelium as well as atherosclerotic burden, plaque size, and macrophage accumulation in mice. AnxA8 regulates oxLDL-induced adhesion molecules expression in aortic endothelial cells. Our data strongly suggest that AnxA8 promotes disease progression through regulation of adhesion and influx of immune cells to the intima. Endothelial specific silencing of AnxA8 reduced atherosclerosis progression. Therapeutic interventions to reduce AnxA8 expression may delay atherosclerosis progression.
© 2025 The Author(s). Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.

Functional cell death pathways are essential for normal ocular vascular development and tissue homeostasis. As our understanding of necrosis-based cell death pathways has expanded, the inclusion of regulated forms, including necroptosis, ferroptosis, and oxytosis, has occurred. Although the existence of these pathways is well described, our understanding of their role during vascular development and pathological neovascularization is very limited. Here, we examined the role of receptor-interacting protein kinase-3 (Ripk3), a key regulator of necroptosis, in postnatal retinal vascularization and retinal and choroidal neovascularization under pathological conditions. Postnatal vascularization of the retinal superficial layer in the absence of Ripk3 (Ripk3-/-) was not significantly different from wild-type mice. However, we noted decreased retinal endothelial cells and pericyte numbers at 3 weeks of age when the formation of the retinal primary vascular plexus was complete. In contrast, choroidal and retinal neovascularization following laser treatment and oxygen-induced ischemic retinopathy increased in the absence of Ripk3 expression, respectively. In addition, the inhibition of RIPK1/3 activity suppressed choroidal neovascularization. Thus, Ripk3 expression and/or activity may have unique roles during normal and pathological ocular vascularization through its interactions with Caspase 8 and modulation of cell death processes.

Bacillus anthracis causes anthrax through a combination of bacterial infection and toxemia. As a major virulence factor of B. anthracis, anthrax lethal toxin (LT) is a zinc-dependent metalloproteinase, exerting its cytotoxicity through proteolytic cleavage of the mitogen-activated protein kinase kinases, thereby shutting down the MAPK pathways. Anthrax lethal toxin induces host lethality mostly by targeting the cardiovascular system. Although the enzymatic activity and the molecular targets of LT have long been known, the detailed mechanisms underlying cellular/tissue/organ toxicity are still poorly understood. In this work, we sought to investigate the mechanism of LT-induced cellular damage in the cardiovascular system. We demonstrate for the first time that anthrax lethal toxin has potent inhibitory effects on the central metabolism of cardiomyocytes and endothelial cells. This is likely due to the observed downregulating of c-Myc expression through the toxin-induced inhibition of the ERK pathway. Since c-Myc is a master transcription factor controlling the expression of many rate-limiting metabolic enzymes in glycolysis and the tricarboxylic acid cycle, LT's downregulation of c-Myc may lead to the observed bioenergetic collapse, particularly, in cardiomyocytes. Since cardiac cell contraction requires continuous production of large amounts of ATP, potent inhibition of the bioenergetics of cardiomyocytes would be incompatible with life. Thus, LT-induced lethality through targeting cardiomyocytes and endothelial cells appears to be a consequence of a bioenergetic collapse, likely due to the toxin's potent inhibitory activity on the MEK-ERK-c-Myc-metabolic/bioenergetic axis within these target cells of cardiovascular system.IMPORTANCEAnthrax lethal toxin (LT) is a major virulence factor of Bacillus anthracis, the causative pathogen of anthrax disease. Anthrax lethal toxin is a metalloproteinase that cleaves and inactivates MEKs, thereby shutting down MAPK pathways, leading to host mortality primarily through targeting of the cardiovascular system. However, the detailed mechanisms underlying the toxin's cellular and tissue toxicity are still poorly understood. Here, we found that anthrax lethal toxin has potent inhibitory activity on glycolysis and oxidative phosphorylation of cardiomyocytes and endothelial cells. These effects appear to be the consequence of downregulation of c-Myc, a master transcription factor that controls many rate-limiting enzymes of glycolysis and the tricarboxylic acid cycle. With the high demand on energy for cardiac contraction, the potent inhibition of cardiomyocyte metabolism by LT would be incompatible with life. This work provides critical insights into why the cardiovascular system is the major in vivo target of LT-induced lethality.