MicroRNAs (miRs) regulate gene expression at the posttranscriptional level and play crucial roles in vascular integrity. As such, they may have a role in modifying abdominal aortic aneurysm (AAA) ...expansion, the pathophysiological mechanisms of which remain incompletely explored. Here, we investigate the role of miRs in 2 murine models of experimental AAA: the porcine pancreatic elastase (PPE) infusion model in C57BL/6 mice and the AngII infusion model in Apoe-/- mice. AAA development was accompanied by decreased aortic expression of miR-29b, along with increased expression of known miR-29b targets, Col1a1, Col3a1, Col5a1, and Eln, in both models. In vivo administration of locked nucleic acid anti-miR-29b greatly increased collagen expression, leading to an early fibrotic response in the abdominal aortic wall and resulting in a significant reduction in AAA progression over time in both models. In contrast, overexpression of miR-29b using a lentiviral vector led to augmented AAA expansion and significant increase of aortic rupture rate. Cell culture studies identified aortic fibroblasts as the likely vascular cell type mediating the profibrotic effects of miR-29b modulation. A similar pattern of reduced miR-29b expression and increased target gene expression was observed in human AAA tissue samples compared with that in organ donor controls. These data suggest that therapeutic manipulation of miR-29b and its target genes holds promise for limiting AAA disease progression and protecting from rupture.
Identification and treatment of abdominal aortic aneurysm (AAA) remains among the most prominent challenges in vascular medicine. MicroRNAs are crucial regulators of cardiovascular pathology and ...represent possible targets for the inhibition of AAA expansion. We identified microRNA-21 (miR-21) as a key modulator of proliferation and apoptosis of vascular wall smooth muscle cells during development of AAA in two established murine models. In both models (AAA induced by porcine pancreatic elastase or infusion of angiotensin II), miR-21 expression increased as AAA developed. Lentiviral overexpression of miR-21 induced cell proliferation and decreased apoptosis in the aortic wall, with protective effects on aneurysm expansion. miR-21 overexpression substantially decreased expression of the phosphatase and tensin homolog (PTEN) protein, leading to increased phosphorylation and activation of AKT, a component of a pro-proliferative and antiapoptotic pathway. Systemic injection of a locked nucleic acid-modified antagomir targeting miR-21 diminished the pro-proliferative impact of down-regulated PTEN, leading to a marked increase in the size of AAA. Similar results were seen in mice with AAA augmented by nicotine and in human aortic tissue samples from patients undergoing surgical repair of AAA (with more pronounced effects observed in smokers). Modulation of miR-21 expression shows potential as a new therapeutic option to limit AAA expansion and vascular disease progression.
Recent genome wide association studies have identified a number of genes that contribute to the risk for coronary heart disease. One such gene, TCF21, encodes a basic-helix-loop-helix transcription ...factor believed to serve a critical role in the development of epicardial progenitor cells that give rise to coronary artery smooth muscle cells (SMC) and cardiac fibroblasts. Using reporter gene and immunolocalization studies with mouse and human tissues we have found that vascular TCF21 expression in the adult is restricted primarily to adventitial cells associated with coronary arteries and also medial SMC in the proximal aorta of mouse. Genome wide RNA-Seq studies in human coronary artery SMC (HCASMC) with siRNA knockdown found a number of putative TCF21 downstream pathways identified by enrichment of terms related to CAD, including "vascular disease," "disorder of artery," and "occlusion of artery," as well as disease-related cellular functions including "cellular movement" and "cellular growth and proliferation." In vitro studies in HCASMC demonstrated that TCF21 expression promotes proliferation and migration and inhibits SMC lineage marker expression. Detailed in situ expression studies with reporter gene and lineage tracing revealed that vascular wall cells expressing Tcf21 before disease initiation migrate into vascular lesions of ApoE-/- and Ldlr-/- mice. While Tcf21 lineage traced cells are distributed throughout the early lesions, in mature lesions they contribute to the formation of a subcapsular layer of cells, and others become associated with the fibrous cap. The lineage traced fibrous cap cells activate expression of SMC markers and growth factor receptor genes. Taken together, these data suggest that TCF21 may have a role regulating the differentiation state of SMC precursor cells that migrate into vascular lesions and contribute to the fibrous cap and more broadly, in view of the association of this gene with human CAD, provide evidence that these processes may be a mechanism for CAD risk attributable to the vascular wall.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Coronary heart disease (CHD) is the leading cause of mortality in both developed and developing countries worldwide. Genome-wide association studies (GWAS) have now identified 46 independent ...susceptibility loci for CHD, however, the biological and disease-relevant mechanisms for these associations remain elusive. The large-scale meta-analysis of GWAS recently identified in Caucasians a CHD-associated locus at chromosome 6q23.2, a region containing the transcription factor TCF21 gene. TCF21 (Capsulin/Pod1/Epicardin) is a member of the basic-helix-loop-helix (bHLH) transcription factor family, and regulates cell fate decisions and differentiation in the developing coronary vasculature. Herein, we characterize a cis-regulatory mechanism by which the lead polymorphism rs12190287 disrupts an atypical activator protein 1 (AP-1) element, as demonstrated by allele-specific transcriptional regulation, transcription factor binding, and chromatin organization, leading to altered TCF21 expression. Further, this element is shown to mediate signaling through platelet-derived growth factor receptor beta (PDGFR-β) and Wilms tumor 1 (WT1) pathways. A second disease allele identified in East Asians also appears to disrupt an AP-1-like element. Thus, both disease-related growth factor and embryonic signaling pathways may regulate CHD risk through two independent alleles at TCF21.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
OBJECTIVE—Genomewide association studies have implicated allelic variation at 9p21.3 in multiple forms of vascular disease, including atherosclerotic coronary heart disease and abdominal aortic ...aneurysm. As for other genes at 9p21.3, human expression quantitative trait locus studies have associated expression of the tumor suppressor gene CDKN2B with the risk haplotype, but its potential role in vascular pathobiology remains unclear.
METHODS AND RESULTS—Here we used vascular injury models and found that Cdkn2b knockout mice displayed the expected increase in proliferation after injury, but developed reduced neointimal lesions and larger aortic aneurysms. In situ and in vitro studies suggested that these effects were attributable to increased smooth muscle cell apoptosis. Adoptive bone marrow transplant studies confirmed that the observed effects of Cdkn2b were mediated through intrinsic vascular cells and were not dependent on bone marrow–derived inflammatory cells. Mechanistic studies suggested that the observed increase in apoptosis was attributable to a reduction in MDM2 and an increase in p53 signaling, possibly due in part to compensation by other genes at the 9p21.3 locus. Dual inhibition of both Cdkn2b and p53 led to a reversal of the vascular phenotype in each model.
CONCLUSION—These results suggest that reduced CDKN2B expression and increased smooth muscle cell apoptosis may be one mechanism underlying the 9p21.3 association with aneurysmal disease.
Experimental and epidemiological studies suggest that vitamin D may be implicated in haemostatic regulations and influence the risk of venous thromboembolism (VTE). The aim of this study was to ...investigate whether oral supplementation of vitamin D3 combined with calcium reduces the risk of VTE. In the randomised, double-blind, placebo-controlled Women's Health Initiative Calcium Plus Vitamin D trial, 36,282 postmenopausal women aged 50-79 years were randomised to receive 1,000 mg of calcium carbonate and 400 IU of vitamin D3 per day (n=18,176) or a matching placebo (n=18,106) during an average of seven years. This secondary analysis of the trial compared the incidence of VTE by treatment group using an intention-to-treat Cox regression analysis. The incidence of VTE did not differ between women randomised to calcium plus vitamin D and women randomised to placebo (320 vs 348 VTE events, respectively; hazard ratio (HR) 0.92, 95 % confidence interval (CI) 0.79-1.07). Results were not modified in an analysis using inverse-probability weights to take non-adherence into account (HR 0.94, 95 %CI 0.73-1.22) or in multiple subgroups. Whereas the risk of a non-idiopathic VTE was similar between groups, the risk of idiopathic VTE was lower in women randomised to calcium plus vitamin D (40 vs 65 events; HR 0.62, 95 %CI 0.42-0.92). In conclusion, daily supplementation with 1,000 mg of calcium and 400 IU of vitamin D did not reduce the overall incidence of VTE in generally healthy postmenopausal women. However, the observed reduced risk of idiopathic VTE in women randomised to calcium and vitamin D warrants further investigations.
Summary
Experimental and epidemiological studies suggest that vitamin D may be implicated in haemostatic regulations and influence the risk of venous thromboembolism (VTE). The aim of this study was ...to investigate whether oral supplementation of vitamin D3 combined with calcium reduces the risk of VTE. In the randomised, double-blind, placebo-controlled Women’s Health Initiative Calcium Plus Vitamin D trial, 36,282 postmenopausal women aged 50-79 years were randomised to receive 1,000 mg of calcium carbonate and 400 IU of vitamin D3 per day (n=18,176) or a matching placebo (n=18,106) during an average of seven years. This secondary analysis of the trial compared the incidence of VTE by treatment group using an intention-to-treat Cox regression analysis. The incidence of VTE did not differ between women randomised to calcium plus vitamin D and women randomised to placebo (320 vs 348 VTE events, respectively; hazard ratio (HR) 0.92, 95 % confidence interval (CI) 0.79-1.07). Results were not modified in an analysis using inverse-probability weights to take non-adherence into account (HR 0.94, 95 %CI 0.73-1.22) or in multiple subgroups. Whereas the risk of a non-idiopathic VTE was similar between groups, the risk of idiopathic VTE was lower in women randomised to calcium plus vitamin D (40 vs 65 events; HR 0.62, 95 %CI 0.42-0.92). In conclusion, daily supplementation with 1,000 mg of calcium and 400 IU of vitamin D did not reduce the overall incidence of VTE in generally healthy postmenopausal women. However, the observed reduced risk of idiopathic VTE in women randomised to calcium and vitamin D warrants further investigations.
Trial Registration: NCT00000611 (
http://www.clinicaltrials.gov
).
Abstract only Recent genome-wide association studies have identified novel loci associated with coronary heart disease (CHD). One locus identified through the large-scale Coronary ARtery DIsease ...Genome wide Replication And Meta-analysis (CARDIoGRAM) study is located on chromosome 6 at q23.2, within the locus encoding the basic helix-loop-helix (bHLH) transcription factor TCF21. This class of transcription factors is well known to regulate cell fate decisions that are critical for embryonic developmental and disease-related pathways, and TCF21 expression has been shown to mark progenitor cells that give rise to the coronary circulation. Six of the most highly associated SNPs in the CARDIoGRAM study are correlated expression quantitative trait loci (eQTL) for TCF21, identifying this as the causal gene. To enable further study of the mechanisms by which allelic variation at 6q23.2 regulates TCF21 expression, we have pursued studies to identify which of the CHD-associated SNPs is causal. The best approach is to investigate allelic-expression imbalance, to compare the relative levels of expression of the two alleles in subjects heterozygous for SNPs in the coding region and then relating any observed imbalance to the genotype at associated SNPs. Peripheral blood samples from the GenePAD study have been used for the isolation of both genomic DNA and white blood cell total RNA, and allelic imbalance determined by Taqman genotyping assays at rs12190287. We identified a significant difference in allele expression in 13 heterozygous Caucasian subjects, with a normalized mean Taqman CT value from the disease major allele (C) to be 2.35, and 1.94 for the minor protective allele (G), P <0.0001. These data are consistent with disease association related to decreased TCF21 expression, and are consistent with rs12190287 being the disease-associated variation in this locus.
Abstract only Introduction: Recent genome-wide association studies have identified a number of genes that contribute to the risk for coronary heart disease, including TCF21, a bHLH transcription ...factor believed to serve a critical role in the differentiation of epicardial progenitor cells that give rise to coronary smooth muscle. Methods and Results: Results of immunolocalization and in situ hybridization studies showed that TCF21 is not expressed in medial smooth muscle cells (SMC) of adult human coronary arteries but primarily within the adventitia and within cells of unknown origin in atherosclerotic lesions. LacZ reporter animal studies with ApoE null background showed a shift in Tcf21 expression from adventitial and medial cells to the neointima, notably into the area below and within the forming fibrous cap, suggesting that Tcf21 expressing cells migrate into the forming atherosclerotic lesion. To investigate disease-related functions of TCF21 in SMCs, knockdown and overexpression studies were performed in human coronary artery smooth muscle cells, revealing an effect on migration, proliferation and SMC marker gene expression. Genome-wide RNA-seq studies in these cells with siRNA treatment identified putative TCF21 downstream genes and pathways involved in vascular development as well as atherosclerosis. Supporting ChIP-seq data suggests that these - as over half of all differentially expressed genes - are direct transcriptional targets. To elucidate the behavior of Tcf21 expressing cells in the development of disease lesions, we are employing an in vivo Tcf21 lineage tracing model. Preliminary data from these studies suggests that Tcf21 expressing cells give rise to fibrous cap SMCs, and ongoing studies are aimed at the further characterization of molecular pathways that are active in these Tcf21 expressing cells. Conclusions: The gene regulatory and expression profile of Tcf21 during disease progression strongly implies a key role in Coronary Heart Disease. Tcf21 is likely to play a role in the migration of SMC progenitors from the adventitia into the forming atherosclerotic plaque. As a source of differentiated SMCs of the fibrous cap Tcf21 may therefore promote plaque stability and protect from lesion rupture.
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