ANALYSIS OF DIFFERENTIAL ExPRESSION OF MATRIx METALLOPROTEASES IN STABLE AND UNSTABLE ATHEROSCLEROTIC LESIONS BY A METHOD OF FULL GENOME SEQUENCING OF RNA: PILOT STUDY

Abstract

Aim. To analyze differential expression of metalloproteases genes, involved into the processes of stabilization/destabilization of atherosclerotic plaque, with the method of full genome sequencing of RNA, and to evaluate the level of metalloproteases in homogenates of plaques of various types by immune enzyme assay method (IEA).Material and methods. The study has been conducted on the specimens of atherosclerotic plaques of patients aged 45-65 y.o., inhabitants of Western Siberia with angiographically proven coronary atherosclerosis and no acute coronary syndrome, with stable angina II-IV functional class. Specimens collection from the plaques was done during an operation if there were intraoperational indications. Histology performed. In intima/media homogenates by IEA method, with BCM Diagnostics assays the levels of destructive markers were measured: MMP-1, MMP3, MMP-7, MMP-9, TIMP-1 on the Multiscan EX (Thermo Fisher Scientific, USA). Libraries preparation for full genomic sequencing of RNA was done with Illumina’s TruSeq RNA Sample Preparation Kit (Illumina, USA). Expression profile in tissues was done on HiSeq 1500 (Illumina, USA).Results. There are differences in expression of the genes MMP2, MMP7, MMP8, MMP9, MMP12, и MMP14 in different types of plaques. There was 8 times higher significant raise in increase of the expression level of ММР9 (p<0,001) in unstable plaque of dystrophic-necrotic type. Study by IEA of MMP-7 content, which is an activator of pro-MMP-9, as well as the content of MMP-9 itself, showed their increased levels in unstable plaques comparing to fatty streaks (1,5 and 2,4 times) and young stable plaques (1,4 and 2,1 times).Conclusion. For the gene ММР9 there were significant differences obtained, of expression levels in stable atherosclerotic fibrous plaque and unstable plaque of dystrophic-necrotic type. With the IEA it was found that fatty streaks and young stable atheromas of coronary arteries have an increased concentration of MMP-3 and decreased activity of tissue inhibitor of metalloproteases. In unstable plaques with the tendency to rupture/ulceration there are increased levels of MMP-1, MMP-7, MMP-9.

Keywords

• matrix metalloproteases
• atherosclerotic plaques
• rna
• full genomic sequencing of rna
• transcriptome
• atherosclerosis