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Chemotactic cytokines and their receptors play an important role in CVDs. The progress of atherosclerosis may cause MI. MI induces an inflammatory immune response, which causes a progressive expansion of the infarcted area and dilation of the ventricles with replacement of the adjacent myocardium by connective tissue, the so-called "remodeling" of the heart muscle. The regeneration of the blood vessel network system by recruitment of HCs for recovering the supply of the infarcted area with oxygen, nutrients, and the prevention of apoptosis of myocytes can be beneficial for heart function. In order to gain further knowledge in the development of atherosclerosis the role of platelet-CX3CR1 receptor and his CX3CL1 ligand was studied during the formation of PMC. The first part of this study includes the recombinant expression of the Met-CCL5, protease-resistant CXCL12 (S4V) and F1-CX3CL1 in E. coli. The chemokines were expressed with different techniques as inclusion bodies in the cytoplasm and were purified by sequential steps with chromatographic methods using FPLC. The second part of this study includes the design of a novel therapeutic strategy with Met-CCL5, a chemokine receptor antagonist to suppress initial neutrophil infiltration, and a protease-resistant CXCL12 (S4V) variant, which is inert to MMP-2 and exopeptidase cleavage, but retains its chemotactic bioactivity for recruitment of circulating HCs, promoting neovascularization after MI. To control the proper timing and local release of Met-CCL5 and CXCL12 (S4V), two different biodegradable hydrogels were implemented, a FDH for delivering Met-CCL5 over 24 h and a SDH for a gradual release of CXCL12 (S4V) over 4 weeks. In a mouse model, intramyocardial injection of these agents reduced infarct size, blocked neutrophil recruitment into the affected tissue, stimulated capillary formation, and improved cardiac function after MI. Last part of this study showed whether platelet-CX3CR1 plays a role in PMC formation. The platelets were preincubated with the CX3CR1-antagonist F1-CX3CL1 and the PMC formation was evaluated. This study describes the expression of different chemokines which play an important role in CVDs. Furthermore, time-controlled release of Met-CCL5-FDH and CXCL12 (S4V)-SDH could demonstrate blocking of neutrophil recruitment to the inflamed myocardium and recruiting HC from circulating blood, thereby improving cardiac function after MI. As result, this study describes a novel and a promising therapeutic strategy to sustain the endogenous reparatory mechanisms, and may represent a potentially clinical relevant alternative to cell-based therapies. Additionally, the formation of PMC and the detection of platelet-bound CX3CL1 on SMCs suggest a significant involvement of the CX3CR1–CX3CL1 axis in platelet accumulation and monocyte recruitment at sites of arterial injury in atherosclerosis. The F1-CX3CL1 antagonist inhibits the formation of the interaction of the CX3CR1–CX3CL1 axis and the formation of PMC is reduced. more...

Embo molecular medicine 5(3), 471-481 (2013). doi:10.1002/emmm.201201717, Published by Wiley-Blackwell, Chichester