P387Secretome modulation by high-cholesterol diet in pig coronary arteries

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Abstract

Purpose

Atherosclerosis connected diseases are the leading cause of death in Europe, accounting for over two million deaths each year. A high-cholesterol level is considered an important risk factor in the development of atherosclerosis, and ischemic heart disease. In the present study an animal model was exploited in order to obtain insights on both plaque induction and its progression. For this purpose, proteins which are actively synthesized and secreted or released (secretome) by coronaries isolated from pig fed with hypercholesterolemic diet were identified and compared with secretion by normal coronary arteries.

Methods

Coronaries isolated from pigs eating standard (SD) or high cholesterol diet (HCD) were incubated for 24 hrs in serum-free medium. Released proteins were digested with trypsin. Peptide mixtures were fractioned by nano-HPLC and analyzed by mass spectrometry. Proteins reported as differentially modulated were statistical validated across 6 replicates (T-test > 95%; p value < 0.05). Histological characterization on SD and HCD coronary vessels was performed.

Results

With our proteomics investigation we were able to identify about 100 released proteins and/or degradation products of cellular proteins. Among these, 13 proteins resulted differentially modulated between control and high cholesterol diet. Several modulated proteins are intracellular (i.e myosin) and among these, some are involved in the adhesion pathway formation (i.e. vinculin). Furthermore, the extracellular matrix components (i.e. perlecan) resulted over-expressed in HCD samples suggesting an important and increased proteolytic activity. The expression in the artery wall of the most interesting factors was documented by immunofluorescence.

Conclusions

Secretome by coronary arteries with early plaques in HCD was able to identify specific, modulated factors and molecular pathways. The proteomic strategy could help to disclose molecular components involved in atherosclerosis onset and development, with the possibility of their detection in peripheral blood as circulating substances.

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