Frauke S. Czepluch

Leptin-Dependent and Leptin-Independent Paracrine Effects of Perivascular Adipose Tissue on Neointima Formation

Objective—Clinical and experimental evidence suggests that periadventitial adipose tissue may modulate vascular lesion formation. The aim of this study was to determine the role of perivascular leptin expression on neointima formation and to differentiate it from local inflammation and systemically elevated leptin levels. Approach and Results—Increased neointima formation after carotid artery injury was observed in hyperleptinemic, diet–induced obese wild-type mice, but not in leptin-deficient ob/ob mice. High-fat diet was associated with increased leptin expression in visceral adipose tissue (VAT) as well as in perivascular adipose tissue. Perivascular leptin overexpression achieved by adenoviral vectors enhanced intimal cell proliferation and neointima formation in wild-type mice, but not in leptin receptor–deficient mice. Perivascular transplantation of VAT from high-fat diet–induced obese wild-type mice around the carotid artery of immunodeficient mice also promoted neointima formation, without affecting body weight or systemic leptin levels, and this effect was absent, if VAT from ob/ob mice was used. On the contrary, perivascular transplantation of VAT from ob/ob mice fed high-fat diet, characterized by marked immune cell accumulation, promoted neointimal hyperplasia also in the absence of leptin. In vitro, recombinant leptin and VAT-conditioned medium increased human arterial smooth muscle cell proliferation in a (partly) leptin-dependent manner. Conclusions—Our findings suggest that locally elevated leptin levels may promote neointima formation, independent of obesity and systemic hyperleptinemia, but also underline the importance of perivascular inflammation in mediating the increased cardiovascular risk in obesity.

Increased proatherogenic monocyte-platelet cross-talk in monocyte subpopulations of patients with stable coronary artery disease.

Monocytes and platelets are important cellular mediators of atherosclerosis. Human monocytes can be divided into CD14++CD16−, CD14++CD16+ and CD14+CD16++ cells, which differ in their functional properties. The aim of this study was to examine monocyte subset distribution, monocyte–platelet aggregate (MPA) formation and expression of CCR5, the receptor of the platelet‐derived chemokine CCL5, and to determine whether these parameters are altered in individuals with coronary atherosclerosis.

In Vitro and In Vivo Effects of Human Monocytes and their Subsets on New Vessel Formation

Human monocytes can be divided into CD16− monocytes and CD16+ monocytes. Studies in mice suggested differential effects of monocyte subsets during new vessel formation.

Stage-dependent detection of CD14+ and CD16+ cells in the human heart after myocardial infarction

Monocytes are critically involved in cardiovascular wound healing processes. Human monocytes can be classified into two subsets based on the expression of CD14 and CD16. Here, we examined the temporal and spatial distribution of CD14+ and CD16+ cells after myocardial infarction (MI) in human heart and spleen tissue and correlated it with markers of cardiac repair. Heart samples obtained at autopsy were histologically classified into acute (AMI; n = 11), subacute (SAMI; n = 10) and old (OMI; n = 16) MI, or control myocardium (CONTR; n = 8). Histochemical analyses revealed marked fibrosis in OMI (p < 0.001 vs. CONTR). The adhesion molecule CD56 was also strongly expressed in OMI (p < 0.01 vs. CONTR) and found to correlate with fibrosis (p < 0.001). The number of capillaries was reduced in OMI (p < 0.01 vs. CONTR; p < 0.05 vs. AMI), whereas the hypoxia indicator carbonic anhydrase IX was predominantly expressed in AMI (p < 0.01 vs. OMI and CONTR) and SAMI (p < 0.05 vs. OMI and CONTR). The monocyte chemoattractrant osteopontin was also more highly expressed in hearts of SAMI patients (p < 0.01 vs. CONTR). Numbers of CD14+ monocytes were found to correlate with CD16+ cells (p < 0.05) and inversely with fibrosis (p < 0.05). Regarding a MI-associated release of monocytes from spleen reservoirs, a non-significant reduction of splenic CD14+ and CD16+ cells was detected in subjects with AMI. In conclusion, disease stage-specific alterations in CD14+ and CD16+ cells in human heart may contribute to cardiac repair processes following MI.

Circulating endothelial cells expressing the angiogenic transcription factor Krüppel‐like factor 4 are decreased in patients with coronary artery disease

The zinc finger transcription factor KLF4 is known to control diverse EC functions.

Genetische Diagnostik bei Kardiomyopathien

Cardiomyopathies often have a genetic etiology. New genetic diagnostic strategies based on next generation sequencing (NGS)-approaches will continuously increase our knowledge about the genetic basis of cardiomyopathies within the following years. Diagnostics and therapy of rare, genetically-induced cardiac diseases increasingly require special cardiac and genetic knowledge. Interestingly, mutations in the same gene or even identical gene mutations can be associated with different cardiomyopathy phenotypes and can exhibit incomplete penetrance or variable expressivity. In the future, the correct interpretation and classification of novel gene variants identified in patients with inherited cardiomyopathy forms will represent a great challenge. Genetic counselling and - if appropriate - subsequent genetic testing for cardiomyopathy patients and their asymptomatic relatives is essential for an early diagnosis of the disease, a prognostic evaluation and possibly for the start of preventive or therapeutic measures.

Genetic determinants of heart failure: facts and numbers

The relevance of gene mutations leading to heart diseases and hence heart failure has become evident. The risk for and the course of heart failure depends on genomic variants and mutations underlying the so‐called genetic predisposition. Genetic contribution to heart failure is highly heterogenous and complex. For any patient with a likely inherited heart failure syndrome, genetic counselling is recommended and important. In the last few years, novel sequencing technologies (named next‐generation sequencing – NGS) have dramatically improved the availability of molecular testing, the efficiency of genetic analyses, and moreover reduced the cost for genetic testing. Due to this development, genetic testing has become increasingly accessible and NGS‐based sequencing is now applied in clinical routine diagnostics. One of the most common reasons of heart failure are cardiomyopathies such as the dilated or the hypertrophic cardiomyopathy. Nearly 100 disease‐associated genes have been identified for cardiomyopathies. The knowledge of a pathogenic mutation can be used for genetic counselling, risk and prognosis determination, therapy guidance and hence for a more effective treatment. Besides, family cascade screening for a known familial, pathogenic mutation can lead to an early diagnosis in affected individuals. At that timepoint, a preventative intervention could be used to avoid or delay disease onset or delay disease progression. Understanding the cellular basis of genetic heart failure syndromes in more detail may provide new insights into the molecular biology of physiological and impaired cardiac (cell) function. As our understanding of the molecular and genetic pathophysiology of heart failure will increase, this might help to identify novel therapeutic targets and may lead to the development of new and specific treatment options in patients with heart failure.

CCL5 deficiency reduces neointima formation following arterial injury and thrombosis in apolipoprotein E-deficient mice

Activated platelets secrete different chemokines, among others CCL5, thereby triggering inflammatory cell recruitment into the vessel wall. Here, we investigated how CCL5 deficiency influences vascular remodeling processes. Experiments were performed in apolipoprotein E and CCL5 double deficient (ApoE(-/-)×CCL5(-/-)) mice, using ApoE(-/-)×CCL5(+/+) mice as controls. The ferric chloride model was applied to induce thrombosis at the site of carotid artery injury within minutes and the formation of a smooth muscle cell-rich neointima within 3weeks. In both groups, vascular injury resulted in thrombus formation. CCL5 deficiency did not alter thrombus resolution examined at day 7. Analysis at 21days revealed that CCL5 absence was associated with a significant reduction in the neointima area (p<0.05), neointima-to-media ratio (p<0.05) and lumen stenosis (p<0.05) compared to ApoE(-/-)×CCL5(+/+) mice. Immunohistochemical analysis of CCL5 receptors showed decreased CCR5 positive staining in ApoE(-/-)×CCL5(-/-) mice (p<0.01), whereas the amount of CCR1 (p=0.053) and Mac2-positive macrophages (p<0.05) was increased. The amount of SMA-positive smooth muscle cells was lower in ApoE(-/-) mice lacking CCL5 (p<0.05). Positive staining for Krüppel-like factor 4 (KLF4), an atheroprotective transcription factor, was increased in the neointima of ApoE(-/-)×CCL5(-/-) mice (p<0.05) and found to co-localize with smooth muscle cells. In summary, CCL5 deficiency resulted in reduced neointima formation after carotid artery injury and thrombosis. Hemodynamic and histochemical analyses suggested that this was not due to differences in thrombus formation or resolution. Possibly, the atheroprotective effect of CCL5 deficiency is mediated by KLF4 upregulation in smooth muscle cells.