Robert Stöhr

MicroRNAs in vascular aging and atherosclerosis.

Lipid dysfunction, inflammation, immune response and advanced aging are major factors involved in the initiation and progression of atherosclerosis. MicroRNAs (miRNAs) have emerged as important regulators of gene expression that post transcriptionally modify cellular responses and function. MiRNAs are crucially involved in several vascular pathologies which show a clear association with increasing age (Dimmeler and Nicotera, 2013). Several studies have demonstrated that miRNA dysregulation has a crucial role in the development of atherosclerotic disease, encompassing every step from plaque formation to destabilization and rupture. This review will present the recent advances in the elucidation of the complex pathophysiological mechanisms in vascular aging by which miRNAs regulate the different phases of atherosclerotic process with a focus on endothelial cells and both, innate and adaptive immune systems. Furthermore, the future areas of research and potential clinical strategies will be discussed.

Insulin resistance and atherosclerosis: convergence between metabolic pathways and inflammatory nodes.

For some time now it has been known that diabetes and atherosclerosis are chronic inflammatory diseases that are closely associated with one another and often develop together. In both there is an increase in tissue-wide inflammation that is exhibited by the infiltration of immune cells into the adipose tissue and the vascular walls respectively. The monocyte/macrophage populations that are recruited in these seemingly different settings also display a high similarity by exhibiting similar phenotypes in both conditions. In the insulin resistant as well as the atherosclerotic setting there is a distinct switch in the macrophage populations present from an anti-inflammatory (M2) population to an inflammatory (M1) population, which releases cytokines and chemotactic factors with the ability to worsen the local environment and thus aggravate the situation by creating a vicious circle. However, although some discoveries suggest that preventing the development of M1 macrophages reduces inflammation and thereby aggravation of these diseases, there are currently no clear-cut opinions on how to achieve a switch from M2 to M1.

Thirty-day outcome after transcatheter aortic valve implantation compared with surgical valve replacement in patients with high-risk aortic stenosis: a matched comparison.

BackgroundTranscatheter aortic valve implantation (TAVI) has become a therapeutic alternative to surgery for the treatment of severe aortic stenosis in high-surgical risk patients. The aim of this study was to compare 30-day mortality of high-risk patients treated by TAVI versus surgical aortic valve replacement. MethodsA total of 175 patients (60 men; mean age, 80±6 years; Euroscore 21±13%) having undergone TAVI were compared with 175 matched patients (76 men; mean age, 79±3 years; Euroscore 17±9%), which have undergone conventional aortic valve replacement and were deemed to be high-risk patients by the cardiothoracic surgeons. Thirty-day mortality and major adverse events were recorded in both groups. Patients’ characteristics were analyzed for predictors of mortality in the TAVI group. ResultsTwenty-one patients (12%) in the TAVI group and 13 patients (8%) in the surgical group died within 30 days of the procedure (P=0.165). Two patients (1%) in the TAVI group and one patient (0.5%) in the conventional surgery group had a major stroke (P=1.0). Seven patients (4%) in the TAVI group and 25 patients (14%) in the conventional surgery group required dialysis post procedure (P=0.0013). The average length of stay in the intensive care unit was lower in the TAVI group compared with the conventional surgical group (3.3±3.1 vs. 6.6±10.5 days; P<0.001). Age was the only independent predictor of mortality in the TAVI group (odds ratio=1.009; 95% confidence interval: 1.001–1.018 per additional year; P=0.0186) and in the total study population (odds ratio=1.007; 95% confidence interval: 1.001–1.013 per additional year; P=0.0186). ConclusionIn high-surgical risk patients, TAVI can be performed at a mortality risk comparable with conventional surgery with a reduced length of post interventional intensive care unit stay and less need for dialysis.

Loss of TIMP3 exacerbates atherosclerosis in ApoE null mice

BACKGROUND Tissue inhibitor of metalloproteinase 3 (TIMP3) is a stromal protein that inhibits the activity of various proteases and receptors. We have previously shown TIMP3 to be downregulated in metabolic and inflammatory disorders, such as type 2 diabetes mellitus. We have now generated an ApoE(-/-)Timp3(-/-) mouse model in which, through the use of genetics, metabolomics and in-vivo phenotypical analysis we investigated the role of TIMP3 in the development of atherosclerosis. METHODS AND RESULTS En face aorta analysis and aortic root examination showed that ApoE(-/-)Timp3(-/-) mice show increased atherosclerosis with increased infiltration of macrophages into the plaque. Serum concentration of MCP-1 were elevated in the serum of ApoE(-/-)Timp3(-/-) mice coupled with an expansion of the inflammatory (M1) Gr1+ macrophages, both in the circulation and within the aortic tissue. Targeted analysis of metabolites revealed a trend to reduced short chain acylcarnitines. CONCLUSIONS Our study shows that lack of TIMP3 increases inflammation and polarizes macrophages towards a more inflammatory phenotype resulting in increased atherosclerosis.

PDE4 inhibition reduces neointima formation and inhibits VCAM-1 expression and histone methylation in an Epac-dependent manner

Phosphodiesterase 4 (PDE4) activity mediates cAMP-dependent smooth muscle cell (SMC) activation following vascular injury. In this study we have investigated the effects of specific PDE4 inhibition with roflumilast on SMC proliferation and inflammatory activation in vitro and neointima formation following guide wire-induced injury of the femoral artery in mice in vivo. In vitro, roflumilast did not affect SMC proliferation, but diminished TNF-α induced expression of the vascular cell adhesion molecule 1 (VCAM-1). Specific activation of the cAMP effector Epac, but not PKA activation mimicked the effects of roflumilast on VCAM-1 expression. Consistently, the reduction of VCAM-1 expression was rescued following inhibition of Epac. TNF-α induced NFκB p65 translocation and VCAM-1 promoter activity were not altered by roflumilast in SMCs. However, roflumilast treatment and Epac activation repressed the induction of the activating epigenetic histone mark H3K4me2 at the VCAM-1 promoter, while PKA activation showed no effect. Furthermore, HDAC inhibition blocked the inhibitory effect of roflumilast on VCAM-1 expression. Both, roflumilast and Epac activation reduced monocyte adhesion to SMCs in vitro. Finally, roflumilast treatment attenuated femoral artery intima-media ratio by more than 50% after 4weeks. In summary, PDE4 inhibition regulates VCAM-1 through a novel Epac-dependent mechanism, which involves regulatory epigenetic components and reduces neointima formation following vascular injury. PDE4 inhibition and Epac activation might represent novel approaches for the treatment of vascular diseases, including atherosclerosis and in-stent restenosis.

ITCH modulates SIRT6 and SREBP2 to influence lipid metabolism and atherosclerosis in ApoE null mice.

Atherosclerosis is a chronic inflammatory disease characterized by the infiltration of pro-inflammatory macrophages into a lipid-laden plaque. ITCH is an E3 ubiquitin ligase that has been shown to polarize macrophages to an anti-inflammatory phenotype. We therefore investigated the effect of ITCH deficiency on the development of atherosclerosis. ApoE−/−ITCH−/− mice fed a western diet for 12 weeks showed increased circulating M2 macrophages together with a reduction in plaque formation. Bone marrow transplantation recreated the haemopoietic phenotype of increased circulating M2 macrophages but failed to affect plaque development. Intriguingly, the loss of ITCH lead to a reduction in circulating cholesterol levels through interference with nuclear SREBP2 clearance. This resulted in increased LDL reuptake through upregulation of LDL receptor expression. Furthermore, ApoE−/−ITCH−/− mice exhibit reduced hepatic steatosis, increased mitochondrial oxidative capacity and an increased reliance on fatty acids as energy source. We found that ITCH ubiquitinates SIRT6, leading to its breakdown, and thus promoting hepatic lipid infiltration through reduced fatty acid oxidation. The E3 Ubiquitin Ligase ITCH modulates lipid metabolism impacting on atherosclerosis progression independently from effects on myeloid cells polarization through control of SIRT6 and SREBP2 ubiquitination. Thus, modulation of ITCH may provide a target for the treatment of hypercholesterolemia and hyperlipidemia.

TIMP3 interplays with apelin to regulate cardiovascular metabolism in hypercholesterolemic mice.

Objective Tissue inhibitor of metalloproteinase 3 (TIMP3) is an extracellular matrix (ECM) bound protein, which has been shown to be downregulated in human subjects and experimental models with cardiometabolic disorders, including type 2 diabetes mellitus, hypertension and atherosclerosis. The aim of this study was to investigate the effects of TIMP3 on cardiac energy homeostasis during increased metabolic stress conditions. Methods ApoE−/−TIMP3−/− and ApoE−/− mice on a C57BL/6 background were subjected to telemetric ECG analysis and experimental myocardial infarction as models of cardiac stress induction. We used Western blot, qRT-PCR, histology, metabolomics, RNA-sequencing and in vivo phenotypical analysis to investigate the molecular mechanisms of altered cardiac energy metabolism. Results ApoE−/−TIMP3−/− revealed decreased lifespan. Telemetric ECG analysis showed increased arrhythmic episodes, and experimental myocardial infarction by left anterior descending artery (LAD) ligation resulted in increased peri-operative mortality together with increased scar formation, ventricular dilatation and a reduction of cardiac function after 4 weeks in the few survivors. Hearts of ApoE−/−TIMP3−/− exhibited accumulation of neutral lipids when fed a chow diet, which was exacerbated by a high fat, high cholesterol diet. Metabolomics analysis revealed an increase in circulating markers of oxidative stress with a reduction in long chain fatty acids. Using whole heart mRNA sequencing, we identified apelin as a putative modulator of these metabolic defects. Apelin is a regulator of fatty acid oxidation, and we found a reduction in the levels of enzymes involved in fatty acid oxidation in the left ventricle of ApoE−/−TIMP3−/− mice. Injection of apelin restored the hitherto identified metabolic defects of lipid oxidation. Conclusion TIMP3 regulates lipid metabolism as well as oxidative stress response via apelin. These findings therefore suggest that TIMP3 maintains metabolic flexibility in the heart, particularly during episodes of increased cardiac stress.

A Role for Timp3 in Microbiota-Driven Hepatic Steatosis and Metabolic Dysfunction

The effect of gut microbiota on obesity and insulin resistance is now recognized, but the underlying host-dependent mechanisms remain poorly undefined. We find that tissue inhibitor of metalloproteinase 3 knockout (Timp3(-/-)) mice fed a high-fat diet exhibit gut microbiota dysbiosis, an increase in branched chain and aromatic (BCAA) metabolites, liver steatosis, and an increase in circulating soluble IL-6 receptors (sIL6Rs). sIL6Rs can then activate inflammatory cells, such as CD11c(+) cells, which drive metabolic inflammation. Depleting the microbiota through antibiotic treatment significantly improves glucose tolerance, hepatic steatosis, and systemic inflammation, and neutralizing sIL6R signaling reduces inflammation, but only mildly impacts glucose tolerance. Collectively, our results suggest that gut microbiota is the primary driver of the observed metabolic dysfunction, which is mediated, in part, through IL-6 signaling. Our findings also identify an important role for Timp3 in mediating the effect of the microbiota in metabolic diseases.

Glucose-dependent insulinotropic peptide secretion is induced by inflammatory stimuli in an interleukin-1-dependent manner in mice

Recently, glucagon‐like peptide‐1 (GLP‐1) levels have been found to be increased in response to inflammatory stimuli, leading to insulin secretion and prevention of hyperglycaemia during endotoxemia in mice. In the present study, we assess the relevance of the other incretin hormone, glucose‐dependent insulinotropic peptide (GIP), as a regulator of glucose metabolism under inflammatory conditions. We found that lipopolysaccharide (LPS) increased GIP secretion in a time‐ and dose‐dependent manner in C57BL/6J mice. To elucidate the underlying mechanisms, mice were injected with inflammatory cytokines known to be released by LPS. Circulating GIP levels significantly increased in response to interleukin (IL)‐1β but not IL‐6 or tumour necrosis factor (TNF)‐α administration. Using respective knockout mice we found that LPS‐mediated GIP secretion was selectively dependent on IL‐1 signalling. To evaluate the functional relevance of inflammatory GIP secretion we pretreated mice with the GIP‐receptor antagonist (Pro3)GIP. This blunted LPS‐induced TNF‐α and IL‐6 secretion but did not affect LPS‐induced insulin secretion or blood glucose‐lowering. In conclusion, GIP provides a novel link between the immune system and the gut, with proinflammatory‐immune modulatory function but minor glucose regulatory relevance in the context of acute endotoxemia.

Tick-tock: is your cardiometabolic risk on the clock?

Governing a large amount of cellular processes in mammalian cells is a 24-h regulatory mechanism known as the circadian clock. Through the release of neurohormonal factors, the master central clock is able to regulate the otherwise independent peripheral clocks situated in all vital organs. It has recently been shown that forced misalignment of the circadian cycles, often as a consequence of lifestyle factors, is an independent cardiometabolic risk factor and may thus potentially predispose certain groups, such as nightshift workers, to cardiovascular disease. In this review, we will analyse some of the recent advances regarding circadian clock dysfunction and the development of cardiovascular diseases. Finally, we will touch on the developing link between circadian dysfunction and myocardial infarctions.