Objectives:
ESR7 aims to identify and functionally characterize novel genes specific for vascular SMCs and to understand the underlying mechanisms associated with SMCs phenotypic switch in vascular remodeling and calcification. ESR7 will develop a panel of novel SMCs specific markers for detection of atherosclerotic plaque instability and potential targets for prevention.
Methodologies:
Cell cultures: For in vitro studies we will use human and primary rat VSMCs and follow the time-course of gene and protein expression variations by microarrays, qPCR and WBs. By lentiviral transductions we will knock-down or overexpress selected genes of interest and follow the phenotypic changes in cells related to proliferation, migration and contractility. This project will be correlated with results from ESR3, ESR5 and ESR6 for evaluation of VSMCs behavior in those conditions.
Murine arterial injury models: The project involves rat carotid artery balloon injury and mouse carotid artery ligation models that allow for evaluation of vascular remodelling and intimal hyperplasia encompassing both ECs and VSMCs. Time-dependent changes in gene expression after injury will be followed. Morphological and immunohistochemical stainings will be performed in order to characterize the tissues. Results from this part will be exchanged with ESR9.
Tissue expression: The expression pattern of novel VSMCs markers will be studied by immunohistochemistry on human samples representing normal, diseased and vascular tissues with active remodeling: various arteries and veins, carotid and femoral plaques, abdominal aortic aneurysms, as well as restenosis grafts and arteriovenous fistulas and will be fed into ESR8.
Zebrafish knockdowns: Novel VSMCs proteins in vascular patterning will be examined by phenotypic screenings in zebrafish using antisense morpholino and CRISPR technology. For this project we will use GATA1-dsRED/Flk1-GFP zebrafish strain.
