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The chromosome 9p21 (Chr9p21) locus of coronary artery disease has been identified in the first surge of genome-wide association and is the strongest genetic factor of atherosclerosis known today. Chr9p21 encodes the long non-coding RNA (ncRNA) antisense non-coding RNA in the INK4 locus (ANRIL). ANRIL expression is associated with the Chr9p21 genotype and correlated with atherosclerosis severity. Here, we report on the molecular mechanisms through which ANRIL regulates target-genes in trans, leading to increased cell proliferation, increased cell adhesion and decreased apoptosis, which are all essential mechanisms of atherogenesis. Importantly, trans-regulation was dependent on Alu motifs, which marked the promoters of ANRIL target genes and were mirrored in ANRIL RNA transcripts. ANRIL bound Polycomb group proteins that were highly enriched in the proximity of Alu motifs across the genome and were recruited to promoters of target genes upon ANRIL over-expression. The functional relevance of Alu motifs in ANRIL was confirmed by deletion and mutagenesis, reversing trans-regulation and atherogenic cell functions. ANRIL-regulated networks were confirmed in 2280 individuals with and without coronary artery disease and functionally validated in primary cells from patients carrying the Chr9p21 risk allele. Our study provides a molecular mechanism for pro-atherogenic effects of ANRIL at Chr9p21 and suggests a novel role for Alu elements in epigenetic gene regulation by long ncRNAs. [ABSTRACT FROM AUTHOR]

Unlabelled: Background- The nuclear liver X receptor-alpha (LXR-alpha) has been implicated in the regulation of intracellular cholesterol homeostasis, inflammatory response, and atherosclerosis susceptibility. The aim of the present study was to test whether transgenic expression of LXR-alpha might affect these mechanisms and result in a reduction of atherosclerosis., Methods and Results: We generated mice with macrophage overexpression of mouse LXR-alpha, evidenced by significantly elevated expression levels of LXR-target genes (ABCA1, ABCG1) in these cells. For atherosclerosis studies, mice were crossed onto the LDL-receptor deficient background. Plasma lipids and lipoproteins as well as liver triglycerides were not significantly different between transgenic animals and nontransgenic controls. However, lesion area at the brachiocephalic artery (BCA) was significantly reduced (-83%, P=0.02) in male LXR-alpha transgenic mice. This was associated with a significantly increased cholesterol efflux to acceptor-free media (+24%, P=0.002) and ApoA1 containing media (+20%, P<0.0001) as well as reduced lipopolysaccharide (LPS)-induced NO-release from macrophages of transgenic animals, providing a potential mechanism for the reduction of atherosclerosis., Conclusions: Our data show for the first time that transgenic overexpression of LXR-alpha in macrophages has significant antiatherogenic properties. We conclude that overexpression of LXR-alpha in macrophages might be useful as a therapeutic principle for the prevention of atherosclerosis.

Aim: To evaluate the diagnostic potential of cytokeratin-19 (CK-19) mRNA for the detection of disseminated tumor cells in blood, bone marrow and peritoneal lavage in patients with ductal adenocarcinoma of the pancreas., Methods: Sixty-eight patients with pancreatic cancer (n = 37), chronic pancreatitis (n = 16), and non-pancreatic benign surgical diseases (n = 15, control group) were included in the study. Venous blood was taken preoperatively, intraoperatively and at postoperative d 1 and 10. Preoperative bone marrow aspirates and peritoneal lavage taken before mobilization of the tumor were analyzed. All samples were evaluated for disseminated tumor cells by CK-19-specific nested-PCR and quantitative fluorogenic RT-PCR., Results: CK-19 mRNA expression was increased in 24 (64%) blood samples and 11 (30%) of the peritoneal lavage samples in the patients with pancreatic cancer. In 15 (40%) of the patients with pancreatic cancer, disseminated tumor cells were detected in venous blood and bone marrow and/or peritoneal lavage. In the peritoneal lavage, the detection rates were correlated with the tumor size and the tumor differentiation. CK-19 levels were increased in pT3/T4 and moderately/poorly differentiated tumors (G2/G3). Pancreatic cancer patients with at least one CK-19 mRNA-positive sample showed a trend towards shorter survival. Pancreatic cancer patients showed significantly increased detection rates of disseminated tumor cells in blood and peritoneal lavage compared to the controls and the patients with chronic pancreatitis., Conclusion: Disseminated tumor cells can be detected in patients with pancreatic ductal adenocarcinoma by CK-19 fluorogenic RT-PCR. In peritoneal lavage, detection rate is correlated with tumor stage and differentiation. In the clinical use, CK-19 is suitable for the distinction between malignant and benign pancreatic disease in combination with other tumor-specific markers.