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Nearly 98% of the human genome is transcriptionally active but does not encode proteins, reflecting a wide range of non-coding genome with essential regulatory and structural roles. Understanding the endothelial cell function in vascular biology has been pivotal for grasping the repair mechanisms in ischaemic tissue and during cardiovascular diseases. However, only a tiny portion of the human genome's non-coding region has been studied to understand their role in new mechanisms in endothelial cell biology. Herein, we provide an overview of the non-coding RNAs in endothelium. We also discuss the latest research focusing on canonical and non-canonical aspects in non-coding RNA biology, non-coding RNA, cell-to-cell communication, and transcriptional and epigenetic regulation by non-coding RNA in the endothelium.

Leading a research group as an early career researcher (ECR) in academia presents many challenges. First, it imposes many additional pressures on individuals, causing fear of missing out on a great opportunity that could advance your career. Together, the unsettling nature of short-term or temporary contracts, lack of guidance and the imposter syndrome can trigger a crisis in future leadership. Most leadership positions at universities are held by senior colleagues. ECRs have modest input in decision-making, due to a requirement for specific leadership training and experience with oversight that precedes suitable decision-making. The turbulence of the unprecedented world COVID-19 crisis has been felt disproportionally by many researchers, intensely by those with caring responsibilities. In the current academic climate, navigating either between your postdoctoral or fellowship project, leading others, taking strategic project directions, mentoring or networking may feel like too much. This editorial expresses views on the current state of the matter in academia with suggestions for helpful strategies to employ to meet research endpoints. It also addresses some challenges that new principal investigators and academic leaders may face due to external or institutional change, and provides some tangible advice with action points.

SummaryEnhancer of Zeste Homologue 2 (EZH2) modulates gene transcription during endothelial cell (EC) dysfunction, via interaction with non-coding RNAs (ncRNAs). Thus, EZH2 can act as a rheostat in deposition of histone H3K27 trimethylation (H3K27me3) to repress many genes. We profiled EZH2-RNA interactions using formaldehyde/UV assisted cross-linking ligation and sequencing of hybrids (FLASH-seq) in primary human ECs. Transcriptome-wide EZH2-associated ncRNAs and RNA–RNA interactome were obtained. This approach revealed EZH2 directly binding maternally expressed gene (MEG3) and MEG3:MEG3 hybrid structures. By chromatin immunoprecipitation with sequencing (ChIP-seq) following depletion of MEG3, we discovered that MEG3 targets and controls recruitment of EZH2/H3K27me3 onto a regulatory region of integrin subunit alpha 4 (ITGA4). MEG3 knockdown or pharmacological inhibition of EZH2 de-repressed ITGA4, whilst improving endothelial cell function in vitro, and increasing ITGA4 expression in vivo. Our study demonstrates new role for MEG3, as instrumental in epigenetic regulation of EC function by EZH2, through targeting of integrin-dependent signalling.

Autophagy is an essential cellular quality control process that has emerged as a critical one for vascular homeostasis. Here, we show that trichoplein (TCHP) links autophagy with endothelial cell (EC) function. TCHP localizes to centriolar satellites, where it binds and stabilizes PCM1. Loss of TCHP leads to delocalization and proteasome‐dependent degradation of PCM1, further resulting in degradation of PCM1's binding partner GABARAP. Autophagic flux under basal conditions is impaired in THCP‐depleted ECs, and SQSTM1/p62 (p62) accumulates. We further show that TCHP promotes autophagosome maturation and efficient clearance of p62 within lysosomes, without affecting their degradative capacity. Reduced TCHP and high p62 levels are detected in primary ECs from patients with coronary artery disease. This phenotype correlates with impaired EC function and can be ameliorated by NF‐κB inhibition. Moreover, Tchp knock‐out mice accumulate of p62 in the heart and cardiac vessels correlating with reduced cardiac vascularization. Taken together, our data reveal that TCHP regulates endothelial cell function via an autophagy‐mediated mechanism., The interaction of TCHP with PCM1 regulates basal autophagy through controlling GABARAP stability. Loss of TCHP function results in impaired autophagy, the accumulation of SQSTM1/p62 and endothelial dysfunction.

Introduction MicroRNAs (miRNAs) are small non-coding RNAs that orchestrate genetic networks by modulating simultaneous gene expression and regulates vascular function. Despite recent accumulation of experimental data and the emergence of functional models, the complexity of miRNA-based regulation is still far from being well understood. By performing high-content phenotypic screening (HCS), miR-26b emerged from the screen as a top candidate for ECs proliferation. The overexpression of miR-26b significantly increased EC proliferation, migration and survival. Furthermore, miR-26b mimics increased EC tube formation and branching morphogenesis. HCS and bioinformatic analysis identified both Phosphatase and Tensin homolog (PTEN), a gene critical in cell survival and Bone Morphogenetic Protein Receptor Type 1B (BMPR1B; ALK6), important in calcium homoeostasis, as direct targets of miR-26b. Because BMPR1B was identified as a direct target of mir-26b, we decided to study the role of miR-26b in vessel calcification. To test our hypothesis, miR-26b CRISPR/Cas9 global knock-out (KO) mice were generated. Objective The aim of this study is to characterize how miR-26b can influence vascular calcification and characterize ALK6 as mir-26b target. Methods In this study, we have used HCS and miR-26b KO mice and artery from patients with critical limb ischaemia to prove our hypothesis Results We confirmed that miR-26b was knocked down in the miR-26b KO mice without effecting the expression of other miR-26 family members and the host gene. To test whether miR-26b KO mice were more susceptible to calcification, we extracted the heart and aorta from 3-month-old mice and cultured for 6 days in a high phosphate media with and without warfarin. Histological analysis with the calcium stains von Kossa and alizarin red confirmed that miR-26b KO mice displayed a higher level and susceptibility to calcifying compared to control mice. Expression of the markers of calcification (RUNX2 & ALK6) supported the histology. Immunohistochemistry and qPCR confirmed that the expression of ALK6 was also increased in the miR-26b KO. To further characterise this phenotype, mice were aged and multiple tissues including the aorta were harvested from 3 and 6 month old mice. The aorta and hearts were then prepared for high resolution PET/CT using the highly calcium specific sodium fluoride (18F) probe. This probe allows for the detection of both micro and macro calcification. PET/CT supported our ex-vivo culture model and confirmed that 6-month-old miR-26b KO mice had a higher level of micro-calcification compared to control mice. To investigate the clinical relevance of miR-26b, we analysed artery and muscles from below the knee amputated limbs from patients with critical limb ischaemia. The levels of miR-26b was decreased in the muscle biopsy from the less perfused section of the amputated leg compared to more perfused counterpart. Moreover, miR-26b was downregulated in the calcified tissue portion of the artery in comparison to the soft tissue. Conclusion Taken together these findings demonstrate an important role for miR-26b in vessel calcification and suggest its effects are through the upregulation of BMPR1B; Alk6. Conflict of interest None

Autophagy is an essential cellular quality control process that emerged critical for vascular homeostasis. Here we describe, the role for Trichoplein (TCHP) protein in linking autophagy with endothelial cells (ECs) function. The depletion of TCHP in ECs impairs migration and sprouting. TCHP directly binds PCM1, to regulate degradation of GABARAP, thus leading to a defective autophagy. Mechanistically, TCHP is indispensable for autophagosome maturation and its depletion resulted in the accumulation of SQSTM1/p62 (p62) and unfolded protein aggregates in ECs. The latter process is coupled to TCHP-mediated NF-kB activation. Of note, low levels of TCHP and high p62 levels were detected in primary ECs from patients with coronary artery disease. In addition, Tchp knock-out mice showed accumulation of p62 in the heart and cardiac vessels and reduced cardiac vascularization. Here, we reveal an autophagy-mediated mechanism for TCHP down-regulation, which poses a plausible target for regulation of endothelial function.

MicroRNAs regulate endothelial function and angiogenesis, but their implication in pericyte biology remains undetermined. A PCR array, covering a panel of 379 human microRNAs, showed microRNA-532-5p to be one of the most differentially modulated by hypoxia, which was confirmed by qPCR in both skeletal muscle and adventitial pericytes. Furthermore, microRNA-532-5p was upregulated in murine muscular pericytes early after experimentally induced ischemia, decreasing below baseline after reperfusion. Transfection of human pericytes with anti-microRNA, microRNA-mimic, or controls indicates microRNA-532-5p modulates pro-angiogenic activity via transcriptional regulation of angiopoietin-1. Tie-2 blockade abrogated the ability of microRNA-532-5p-overexpressing pericytes to promote endothelial network formation in vitro. However, angiopoietin-1 is not a direct target of microRNA-532-5p. In silico analysis of microRNA-532-5p inhibitory targets associated with angiopoietin-1 transcription indicated three potential candidates, BACH1, HIF1AN, and EGLN1. Binding of microRNA-532-5p to the BACH1 3′ UTR was confirmed by luciferase assay. MicroRNA-532-5p silencing increased BACH1, while a microRNA-532-5p mimic decreased expression. Silencing of BACH1 modulated angiopoietin-1 gene and protein expression. ChIP confirmed BACH1 transcriptional regulation of angiopoietin-1 promoter. Finally, microRNA-532-5p overexpression increased pericyte coverage in an in vivo Matrigel assay, suggesting its role in vascular maturation. This study provides a new mechanistic understanding of the transcriptional program orchestrating angiopoietin-1/Tie-2 signaling in human pericytes., Graphical Abstract, Angiopoietin-1 is an important regulator of angiogenesis, yet little is known regarding its transcriptional regulation. Slater et al. (2018) demonstrate angiopoietin-1 induction in pericytes occurs via miR-532-5p regulation of BACH1 and modulation of this pathway promotes changes in endothelial permeability, vascular stability, and angiogenesis.

Epigenetic mechanisms may regulate the expression of pro-angiogenic genes, thus affecting reparative angiogenesis in ischemic limbs. The enhancer of zest homolog-2 (EZH2) induces thtrimethylation of lysine 27 on histone H3 (H3K27me3), which represses gene transcription. We explored (i) if EZH2 expression is regulated by hypoxia and ischemia; (ii) the impact of EZH2 on the expression of two pro-angiogenic genes: eNOS and BDNF; (iii) the functional effect of EZH2 inhibition on cultured endothelial cells (ECs); (iv) the therapeutic potential of EZH2 inhibition in a mouse model of limb ischemia (LI). EZH2 expression was increased in cultured ECs exposed to hypoxia (control: normoxia) and in ECs extracted from mouse ischemic limb muscles (control: absence of ischemia). EZH2 increased the H3K27me3 abundance onto regulatory regions of eNOS and BDNF promoters. In vitro RNA silencing or pharmacological inhibition by 3-deazaneplanocin (DZNep) of EZH2 increased eNOS and BDNF mRNA and protein levels and enhanced functional capacities (migration, angiogenesis) of ECs under either normoxia or hypoxia. In mice with experimentally induced LI, DZNep increased angiogenesis in ischaemic muscles, the circulating levels of pro-angiogenic hematopoietic cells and blood flow recovery. Targeting EZH2 for inhibition may open new therapeutic avenues for patients with limb ischemia.

Graphical abstract, 11β-Hydroxysteroid dehydrogenase 1 (11βHSD1; EC 1.1.1.146) generates active glucocorticoids from inert 11-keto metabolites. However, it can also metabolize alternative substrates, including 7β-hydroxy- and 7-keto-cholesterol (7βOHC, 7KC). This has been demonstrated in vitro but its consequences in vivo are uncertain. We used genetically modified mice to investigate the contribution of 11βHSD1 to the balance of circulating levels of 7KC and 7βOHC in vivo, and dissected in vitro the kinetics of the interactions between oxysterols and glucocorticoids for metabolism by the mouse enzyme. Circulating levels of 7KC and 7βOHC in mice were 91.3 ± 22.3 and 22.6 ± 5.7 nM respectively, increasing to 1240 ± 220 and 406 ± 39 nM in ApoE−/− mice receiving atherogenic western diet. Disruption of 11βHSD1 in mice increased (p

11β-Hydroxysteroid dehydrogenase type-1 (11β-HSD1) converts inert cortisone into active cortisol, amplifying intracellular glucocorticoid action. 11β-HSD1 deficiency improves cardiovascular risk factors in obesity but exacerbates acute inflammation. To determine the effects of 11β-HSD1 deficiency on atherosclerosis and its inflammation, atherosclerosis-prone apolipoprotein E-knockout (ApoE-KO) mice were treated with a selective 11β-HSD1 inhibitor or crossed with 11β-HSD1-KO mice to generate double knockouts (DKOs) and challenged with an atherogenic Western diet. 11β-HSD1 inhibition or deficiency attenuated atherosclerosis (74–76%) without deleterious effects on plaque structure. This occurred without affecting plasma lipids or glucose, suggesting independence from classical metabolic risk factors. KO plaques were not more inflamed and indeed had 36% less T-cell infiltration, associated with 38% reduced circulating monocyte chemoattractant protein-1 (MCP-1) and 36% lower lesional vascular cell adhesion molecule-1 (VCAM-1). Bone marrow (BM) cells are key to the atheroprotection, since transplantation of DKO BM to irradiated ApoE-KO mice reduced atherosclerosis by 51%. 11β-HSD1-null macrophages show 76% enhanced cholesterol ester export. Thus, 11β-HSD1 deficiency reduces atherosclerosis without exaggerated lesional inflammation independent of metabolic risk factors. Selective 11β-HSD1 inhibitors promise novel antiatherosclerosis effects over and above their benefits for metabolic risk factors via effects on BM cells, plausibly macrophages.—Kipari, T., Hadoke, P. W. F., Iqbal, J., Man, T. Y., Miller, E., Coutinho, A. E., Zhang, Z., Sullivan, K. M., Mitic, T., Livingstone, D. E. W., Schrecker, C., Samuel, K., White, C. I., Bouhlel, M. A., Chinetti-Gbaguidi, G., Staels, B., Andrew, R., Walker, B. R., Savill, J. S., Chapman, K. E., Seckl, J. R. 11β-hydroxysteroid dehydrogenase type 1 deficiency in bone marrow-derived cells reduces atherosclerosis.

The aging process worsens the human body functions at multiple levels, thus causing its gradual decrease to resist stress, damage, and disease. Besides changes in gene expression and metabolic control, the aging rate has been associated with the production of high levels of Reactive Oxygen Species (ROS) and/or Reactive Nitrosative Species (RNS). Specific increases of ROS level have been demonstrated as potentially critical for induction and maintenance of cell senescence process. Causal connection between ROS, aging, age-related pathologies, and cell senescence is studied intensely. Senescent cells have been proposed as a target for interventions to delay the aging and its related diseases or to improve the diseases treatment. Therapeutic interventions towards senescent cells might allow restoring the health and curing the diseases that share basal processes, rather than curing each disease in separate and symptomatic way. Here, we review observations on ROS ability of inducing cell senescence through novel mechanisms that underpin aging processes. Particular emphasis is addressed to the novel mechanisms of ROS involvement in epigenetic regulation of cell senescence and aging, with the aim to individuate specific pathways, which might promote healthy lifespan and improve aging.

Vascular dysfunction is a common consequence of diabetes mellitus. Stable propagation of gene expression from cell to cell generation during development of diseases (like diabetes) is regulated by epigenetic mechanisms. These are heritable patterns of gene expression that cannot solely be explained by changes in DNA sequence. Recent evidence shows that diabetes-induced epigenetic changes can affect gene expression in vascular endothelial cells and vascular smooth muscles cells. Such effects further influence inflammatory and insulin production pathways in these cells and thus ensure a long-term memory, whereby epigenetic changes are maintained even long after restoring normo-glycaemic conditions by appropriate therapeutic approaches. This review focuses on the epigenetic marks, which endure on the vascular chromatin under diabetic conditions.

Objective— The p75 neurotrophin receptor (p75 NTR ) contributes to diabetes mellitus−induced defective postischemic neovascularization. The interleukin-33 receptor ST2 is expressed as transmembrane (ST2L) and soluble (sST2) isoforms. Here, we studied the following: (1) the impact of p75 NTR in the healing of ischemic and diabetic calf wounds; (2) the link between p75 NTR and ST2; and (3) circulating sST2 levels in critical limb ischemia (CLI) patients. Methods and Results— Diabetes mellitus was induced in p75 NTR knockout ( p75KO ) mice and wild-type ( WT ) littermates by streptozotocin. Diabetic and nondiabetic p75KO and WT mice received left limb ischemia induction and a full-thickness wound on the ipsilateral calf. Diabetes mellitus impaired wound closure and angiogenesis and increased ST2 expression in WT , but not in p75KO wounds. In cultured endothelial cells, p75 NTR promoted ST2 (both isoforms) expression through p38 MAPK /activating transcription factor 2 pathway activation. Next, sST2 was measured in the serum of patients with CLI undergoing either revascularization or limb amputation and in the 2 nondiabetic groups (with CLI or nonischemic individuals). Serum sST2 increased in diabetic patients with CLI and was directly associated with higher mortality at 1 year from revascularization. Conclusion— p75 NTR inhibits the healing of ischemic lower limb wounds in diabetes mellitus and promotes ST2 expression. Circulating sST2 predicts mortality in diabetic CLI patients.

Background & Aims: Suppression of the hypothalamic-pituitary-adrenal axis occurs in cirrhosis and cholestasis and is associated with increased concentrations of bile acids. We investigated whether this was mediated through bile acids acting to impair steroid clearance by inhibiting glucocorticoid metabolism by 5 beta-reductase. Methods: The effect of bile acids on glucocorticoid metabolism was studied in vitro in hepatic subcellular fractions and hepatoma cells, allowing quantitation of the kinetics and transcript abundance of 5 beta-reductase. Metabolism was subsequently examined in vivo in rats following dietary manipulation or bile duct ligation. Finally, glucocorticoid metabolism was assessed in humans with obstructive jaundice. Results: In rat hepatic cytosol, chenodeoxycholic acid competitively inhibited 5 beta-reductase (K-i 9.19 +/- 0.40 mu M) and reduced its transcript abundance (in H4iiE cells) and promoter activity (reporter system, HepG2 cells). In Wistar rats, dietary chenodeoxycholic acid (1% w/w chow) inhibited hepatic 5 beta-reductase activity, reduced urinary excretion of 3 alpha,5 beta-tetrahydrocorticosterone and reduced adrenal weight. Conversely, a fat-free diet suppressed bile acid levels and increased hepatic 5 beta-reductase activity, supplementation of the fat-free diet with CDCA reduced 5 beta-reductase activity, and urinary 3 alpha,5 beta-reduced corticosterone. Cholestasis in rats suppressed hepatic 5 beta-reductase activity and transcript abundance. In eight women with obstructive jaundice, relative urinary excretion of 3 alpha,5 beta-tetrahydrocortisol was significantly lower than in healthy controls. Conclusion: These data suggest a novel role for bile acids in inhibiting hepatic glucocorticoid clearance, of sufficient magnitude to suppress hypothalamic-pituitary-adrenal axis activity. Elevated hepatic bile acids may account for adrenal insufficiency in liver disease.

The communication between vascular endothelial cells (ECs) and pericytes in the microvasculature is fundamental for vascular growth and homeostasis; however, these processes are disrupted by diabetes. Here we show that modulation of p75NTR expression in ECs exposed to high glucose activates transcription of miR-503, which negatively affects pericyte function. p75NTR activates NF-κB to bind the miR-503 promoter and upregulate miR-503 expression in ECs. NF-κB further induces activation of Rho kinase and shedding of endothelial microparticles carrying miR-503, which transfer miR-503 from ECs to vascular pericytes. The integrin-mediated uptake of miR-503 in the recipient pericytes reduces expression of EFNB2 and VEGFA, resulting in impaired migration and proliferation. We confirm operation of the above mechanisms in mouse models of diabetes, in which EC-derived miR-503 reduces pericyte coverage of capillaries, increased permeability and impaired post-ischaemic angiogenesis in limb muscles. Collectively, our data demonstrate that miR-503 regulates pericyte–endothelial crosstalk in microvascular diabetic complications., Vascular function and repair is impaired in patients with diabetes. Here, Caporali et al. report that activation of the neurotrophin receptor in vascular endothelial cells induces the antiangiogenic miR-503, which impairs the function of neighbouring pericytes upon microparticle-mediated transfer.

Objective— Gestational diabetes mellitus (GDM) produces fetal hyperglycemia with increased lifelong risks for the exposed offspring of cardiovascular and other diseases. Epigenetic mechanisms induce long-term gene expression changes in response to in utero environmental perturbations. Moreover, microRNAs (miRs) control the function of endothelial cells (ECs) under physiological and pathological conditions and can target the epigenetic machinery. We investigated the functional and expressional effect of GDM on human fetal ECs of the umbilical cord vein (HUVECs). We focused on miR-101 and 1 of its targets, enhancer of zester homolog-2 (EZH2), which trimethylates the lysine 27 of histone 3, thus repressing gene transcription. EZH2 exists as isoforms α and β. Approach and Results— HUVECs were prepared from GDM or healthy pregnancies and tested in apoptosis, migration, and Matrigel assays. GDM-HUVECs demonstrated decreased functional capacities, increased miR-101 expression, and reduced EZH2- β and trimethylation of histone H3 on lysine 27 levels. MiR-101 inhibition increased EZH2 expression and improved GDM-HUVEC function. Healthy HUVECs were exposed to high or normal d -glucose concentration for 48 hours and then tested for miR-101 and EZH2 expression. Similar to GDM, high glucose increased miR-101 expression. Chromatin immunoprecipitation using an antibody for EZH2 followed by polymerase chain reaction analyses for miR-101 gene promoter regions showed that both GDM and high glucose concentration reduced EZH2 binding to the miR-101 locus in HUVECs. Moreover, EZH2-β overexpression inhibited miR-101 promoter activity in HUVECs. Conclusions— GDM impairs HUVEC function via miR-101 upregulation. EZH2 is both a transcriptional inhibitor and a target gene of miR-101 in HUVECs, and it contributes to some of the miR-101-induced defects of GDM-HUVECs.

Purpose: Uncontrolled hyperglycemia is responsible for vascular complications in diabetes. Together with pericytes, endothelial cells (ECs) establish direct cell-to-cell communication in the blood vessel that is fundamental for the regulation of microvascular formation, stabilisation and function. Notably, circulating microparicles (MPs) can serve as signalling molecules and mediators of inter-cellular communication. Recently, we also demonstrated that in ECs microRNA-503 (miR-503) and neurotrophin receptor p75 (p75NTR) individually facilitate diabetes-induced impairment of EC function. This study investigated how expression of miR-503 and its release are regulated by p75NTR in diabetes, and what role do miR-503-carrying microparticles (MPs-miR-503) play in communication between EC and vascular pericytes. Methods: Type 1 diabetes was induced in mice by injection of streptozotocin. Mice underwent unilateral limb ischemia followed by gene transfer using adenoviral vectors. Blood flow recovery was monitored by laser Doppler and post-ischemic angiogenesis was assessed by immunostaining. ECs were cultured in high glucose and reduced of growth factors to mimic diabetes and ischemia. Binding of NF-kB to miR-503 promoter was verified by chromatin immunoprecipitation. MPs were prepared by centrifugation and validated by flow cytometric analysis as events with 0.1-1mm diameter expressing Annexin V. To study the trafficking between ECs and pericytes, an in vitro co-culture system was used, along with an ex-vivo model from diabetic limb muscles. Results: Our findings show that expression of miR-503 and p75NTR increase in ECs in vitro, due to high glucose, as well as in diabetes with ischemia in vivo. Modulation of p75NTR expression regulates transcription of miR-503 and consequently EC function and post-ischemic angiogenesis. We provide evidence that p75NTR promotes activation of NF-kB in ECs, which binds the promoter of miR-503 thus upregulating its expression. NF-kB further induces the shedding of MPs-miR-503 from the ECs, through activation of Rho kinase (ROCK). Circulating MPs-miR-503 are engulfed by vascular pericytes in a RAGE (receptor for advance glycation end product)-dependent manner. This reduces the expression of miR-503 targets like ephrinB2 (EFNB2) and vascular endothelial growth factor-A (VEGFA), and causes functional impairments in pericytes. Conclusion: Here we describe a newly-established mechanism for regulation and release of miRNA-503 into MPs during diabetes-induced vascular diseases.

11β-Hydroxysteroid dehydrogenase type 1 (11β-HSD1) regenerates active glucocorticoids, amplifying intracellular actions.1 11β-HSD1 deficiency or inhibition improves metabolic syndrome and attenuates atherosclerosis in vulnerable rodent strains and is a target for drug development.2–4 However, 11β-HSD1 also catalyses conversion of 7-ketocholesterol,5 which accumulates in fatty tissues,6 to potentially more atherogenic 7β-hydroxycholesterol. Whether atheroprotection with 11β-HSD1 deficiency is dependent on glucocorticoid or oxysterol effects is unknown. Male atherosclerosis-prone ApoE−/− and ApoE−/−.11β-HSD1−/− double knockout (DKO) mice underwent adrenalectomy or sham surgery (n=8/group), then received a high (0.2%) cholesterol Western diet for 12 weeks. The aorta and branches were perfusion-fixed. Lesion volume and extracellular lipids were determined by 3D optical projection tomography. Data are mean±SE of the means. Adrenalectomy had no effect on body/organ weights in either genotype. Removal of endogenous glucocorticoids by adrenalectomy in ApoE−/− mice did not reduce lesion volume (232±24 vs 235±34 μm 3 sham control). DKO mice had reduced lesion volumes (139±17 μm 3 ) compared with ApoE−/−(p 3 ). Adrenalectomised DKO mice had increased extracellular lipids (73.6±2.6 μm 3 ) within the lesion compared with either ApoE−/− adrenalectomised (37.4±5.2 μm 3 ), ApoE−/− sham (42.9±5.5 μm 3 ) or DKO sham (44.2±12 μm 3 ) group. Thus circulating glucocorticoids are necessary for 11β-HSD1 deficiency to attenuate atherosclerosis. However, 11β-HSD1 deficiency increases the lipid content of plaques in the absence of glucocorticoids, perhaps owing to accumulation of 7-ketocholesterol? Consequently, both reactions of 11β-HSD1 may be involved in the effects of the enzyme on atherogenesis.

The atherogenic 7-oxysterols, 7-ketocholesterol (7-KC) and 7β-hydroxycholesterol (7βOHC), can directly impair arterial function. Inter-conversion of 7-KC and 7βOHC has recently been shown as a novel role for the glucocorticoid-metabolizing enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1). Since this enzyme is expressed in vascular smooth muscle cells, we addressed the hypothesis that inter-conversion of 7-KC and 7βOHC by 11β-HSD1 may contribute to regulation of arterial function. Incubation (4–24 h) of aortic rings with either 7-KC (25 μM) or 7βOHC (20 μM) had no effect on endothelium-dependent (acetylcholine) or -independent (sodium nitroprusside) relaxation. In contrast, exposure to 7-KC (but not to 7βOHC) attenuated noradrenaline-induced contraction (Emax) after 4 h (0.78 ± 0.28 vs 0.40 ± 0.08 mN/mm; p, Highlights ► Metabolism of 7-oxysterols by 11β-HSD1 may influence arterial function. ► 11β-HSD1 inter-converted 7-ketocholesterol and 7β-hydroxycholesterol in mouse aorta. ► 7-ketocholesterol, but not 7β-hydroxycholesterol, altered aortic contraction. ► 11β-HSD1-mediated metabolism of 7-oxysterols may influence arterial contraction.