Objectives of the PhD position The PhD position title is “Role of AT2 receptor in intimal calcification” (ESR 14). It has already been shown that the renin-angiotensin system (RAS) is involved in intimal calcification. Infusion of Ang II into ApoE−/− mice enhances calcification processes via AT1 receptors. However, the role of the protective arm of the RAS, including the AT2R, in calcification processes has not yet been studied. Since AT2R stimulation counterbalances most of the AT1R actions, it may have a therapeutic potential for the treatment of intimal calcification. Here we would like to test the hypothesis that AT2 receptor inhibits intimal calcification and to elucidate the underlying molecular mechanisms Methodologies: 1) ApoE−/− mice under high-fat diet will be infused with vehicle or with Ang II and treated either with the AT2R agonist C21, the AT2R antagonist PD123319 or with the AT1 receptor blocker olmesartan. Calcium deposition will be assessed by molecular imaging. Expression of calcification-related genes, pro-inflammatory cytokines, receptor activator of nuclear factor-κB ligand (RANKL) system and angiotensin receptors will be analysed by qPCR and immunohistology. 2) Aortic VSMC will be isolated from WT or AT2R knock out mice, cultured in normal or osteogenic medium. Cells will be stimulated with Ang II or RANKL and treated with C21, PD123319 or with olmesartan. Calcification will be assessed by Alizarin Red staining. Differential gene expression will be analysed by Affymetrix GeneChip® Transcriptome Array, covering both coding and non-coding RNA and validated by qPCR. In addition, it will be investigated whether AT2R stimulation can prevent phenotypic switching of VSMC from contractile to synthetic VSMC. 3) Differentially expressed genes in above-mentioned in vitro experiments will be analysed in samples from in vivo studies. 4) Relevant pathways will be dissected by GO and pathway analysis of microarray data.

Expected results of the PhD research project This project will define the renin-angiotensin system (RAS) as critical and modifiable actor in intimal calcification, with emphasis on the protective RAS arm (here: AT2R) and will elucidate the complex AT2R mediated processes in vitro and in vivo models of calcification. Finally, we will validate this novel AT2R based therapeutic approach in preclinical models of vascular calcification, providing a new therapeutic entry to the treatment of this disease

Please note there are strict *eligibility requirements* which apply to all Marie Skłodowska-Curie Early Stage Researchers. At the time of the appointment: Applicants must not have resided or carried out his/her main activity (work, studies, etc.) in the Netherlands for more than 12 months in the 3 years immediately before appointment under the project; Applicants shall also be in the first four years of their research careers at the time of appointment by the host organisation and have not been awarded a doctoral degree. For more information on Marie Skłodowska-Curie Innovative Training Networks (ITNs), please see: ec.europa.eu/mariecurieactions

Qualifications: Bachelor and Master degrees in Biomedical Sciences or equivalent. An excellent command of the English language is essential. A certification to conduct animal experimentation is desired. Experience: Standard laboratory activities (cell culture, molecular biology) as well as a prior experience with animal work are essentials. Previous experience with immunohistochemistry and confocal microscopy is desirable. Knowledge & skills: Knowledge of cell behaviour, reactivity and phenotype is essential. Knowledge of vascular biology, inflammation and exosomes is desirable. Cell culturing, Western blotting, real time PCR, and ELISA are desirable. Good communication skills and analytical skills are essential. Abilities: Interpreting scientific literature and incorporation into project and understanding the research process are essential. Organising and prioritising own work and organising research within the project timetable are essential. Maintaining accurate and up to date records are essential. Attitude & disposition: Following abilities and dispositions are essential: Flexible and co-operative; Good relationships with staff and to communicate effectively; Self-motivated and hardworking; Enquiring, critical approach to work; Willingness to learn new skills; and Willing to travel and go on secondments. Other circumstances: Willingness to work flexibly in order to achieve project demands and targets as agreed with Supervisor is essential. Able to travel to workshops and for research collaboration in other EU countries with notice is essential.

Secondments for the PhD research project The ESR will rotate in the group of Prof. Marx at Universitätsklinikum Aachen (Germany) for 9 months in order to perform experiments on phenotypic switching of VSMC from contractile to synthetic VSMC. Furthermore, the ESR will stay for 1 month with Vicore Pharma, Göteborg (Per Janssen, Björn Dahlöf), to learn about strategies of drug development, and with a Vicore-associated group of medicinal chemists at Uppsala University (Anders Hallberg, Mats Larhed) to participate in modern drug design of AT2R agonists for 3 months.

Fixed-term contract: 3 years.

The terms of employment of Maastricht University are set out in the Collective Labour Agreement of Dutch Universities (CAO). Furthermore, local UM provisions also apply. For more information look at the website https://www.maastrichtuniversity.nl/support/um-employees

Temporary employment for 3 years.

Your salary would be € 2.222,-- gross per month in the first year up to € 2.840,-- gross per month according to the PhD-student salary scale.

Each year an evaluation will take place.

Maastricht University is renowned for its unique, innovative, problem-based learning system, which is characterized by a small-scale and student-oriented approach. Research at UM is characterized by a multidisciplinary and thematic approach, and is concentrated in research institutes and schools. Maastricht University has around 16,500 students and 4,000 employees. Reflecting the university's strong international profile, a fair amount of both students and staff are from abroad. The university hosts 6 faculties: Faculty of Health, Medicine and Life Sciences, Faculty of Law, School of Business and Economics, Faculty of Humanities and Sciences, Faculty of Arts and Social Sciences, Faculty of Psychology and Neuroscience.

CARIM School for Cardiovascular Diseases

The Cardiovascular Research Institute Maastricht (CARIM), School for Cardiovascular Diseases, is one of the top institutes for translational cardiovascular research in Europe. It is among the world leaders in the fields of research into vascular and thrombotic disorders and atrial fibrillation as well as translational heart failure research. It has also made important international contributions to molecular imaging in the cardiovascular field.

With an annual budget of approximately 22 mln Euros in 2015, CARIM is one of the largest cardiovascular research institutes in Europe, producing more than 550 scientific articles and approximately 45 PhD dissertations per year. CARIM employs 250 researchers from 13 disciplines, about 120 PhD students and 50 technical and support staff members.

Over the last ten years, CARIM papers have been published in high-impact journals such as the New England Journal of Medicine, JAMA, Nature Medicine, Nature Cell Biology, Cell Metabolism, Journal of Experimental Medicine, Circulation, Lancet Neurology, and the Journal of the American College of Cardiology. Bibliometric analyses illustrate that CARIM publications have been cited 1.97 times more often than the world average for the cardiovascular field in the period of 2009 until 2012.

CARIM is recognized by the KNAW as a research school and as an international training site for Early Stage Researchers in the framework of the Marie Curie Program.

Cardiovascular scientists from around the world join CARIM because it values open communication, close cooperation, high ambitions, good facilities and a critical learning environment.