PROJECT 5

Objectives: 
Imbalances between inhibitory and inducing mediators of vascular calcification (VC) lead to an increase in VC. ESR5 will characterize the recently identified inhibitor CBF (“calcification blocking factor”). (1.a) The main focus will be the identification of the active site of the CBF; this will the basis for the development of a peptide mimic for therapeutic application of CBF. (1.b) The impact of this peptide in the context of isolated systolic hypertension will be studied. (2) ESR5 will screen chromatographic fractions from adrenal glands and plasma to identify yet unknown mediators of these processes.

Methodologies: 
1a. Endothelial cells and vascular smooth muscle cells will be cultivated in monoculture as well as in co-culture in the presence of CBF in cooperation with ESR2 (EC) and ESR3 (VSMC), respectively. The resulting shorter peptides of CBF will be identified by FT-mass-spectrometric methods. These peptides will be synthesized (in cooperation with ESR9) and their inhibitory capacity on vascular calcification processes will be evaluated by using an aortic rings. The effect will be compared to the effect of CBF.
1b. ESR will quantify the plasma concentration of patients suffering from isolated systolic hypertension by LC-MS techniques.
2. Bovine adrenal glands and human plasma will be chromatographically fractionated and screened for inhibitory effect of vascular calcification processes by aortic rings. Fraction with inhibitory effects will be identified by mass-spectrometric methods. The anticalcifying activity will be characterized in human aortic smooth muscle cells culture, aortic tissue culture as well as in animal models (VDN rats). Furthermore, the effects on the pulse pressure will be quantified in these animal experiments. The underlying pathways will be identified by RT-PCR analyses of samples the animal experiments (in ESR1). The plasma concentration of the identified inhibitor(s) will be quantified in patients suffering from increased vascular calcification (renal; heart failure) and in corresponding control subjects (in ESR4).