#4536 INFLUENCE OF REDUCTION IN THE MASS OF ACTIVE NEPRONS ON THE EXPRESSION OF MIRNA AND NFκB IN THE MYOCARDIA AND AORT OF WKY RATS

Abstract

Background and Aims At present, there is no doubt about the relationship between the state of the kidneys and the cardiovascular system, which is reflected in the concept of the cardiorenal continuum. The emerging data suggest that miRNAs may be one of the links connecting the kidneys and the cardiovascular system. However, information about the role of epigenomic changes in the development of damage to the heart and blood vessels in chronic kidney disease remains not fully understood. Target. The aim of the work was to evaluate changes in the expression levels of microRNA-21, 133,203, as well as the nuclear transcription factor κB (NFκB) in the tissues of the myocardium and aorta in the development of remodeling of the heart and blood vessels with a chronic reduction in the mass of active nephrons in the experiment. Method Systolic blood pressure (BP, mm Hg), myocardial mass index (MMI, mg/g), relative expression levels of miRNA-21, -133, -203 and NFκB in myocardium and aorta were analyzed in rats Wistar-Kyoto (WKY) with 5/6 nephrectomy (Nx) and duration of experimental exposure 2 mo (n = 9). Controls were sham operated (SO) animals (n = 9). The reverse transcription reaction for ‘complementary’ DNA (cDNA) preparation was carried out under the StemLoop-technology separately for the studied miRNA. The following primers were used in PCR: microRNA-21–5'-GCCCGCTAGCTTATCAGACTGATG-3_, microRNA-133–5_-GCCCGCAGCTGGTAAAATGGAAC-3_, microRNA-203–5_-GCCGGTGAAATGTTTAGGACC-3_ and U6–5_-GCGCGTCGTGAAGCGTTC-3_, and common reverse 5_-GTGCAGGGTCCGAGGT-3. The semi-qualitative evaluation of the miRNA expression level (in relative units, RU) under the 2-(ΔΔCt) protocol at the laboratory referent (0.09) was used in calculations. NFκB expression was quantified using a StemLoop technology and RealTimePCR compared to the expression of the reference-gene GAPDH. Data were analyzed using Student's t-test and the Mann–Whitney, a P<0.05 was considered statistically significant. Results In rats WKY with an experimental decrease of the nephron mass in comparison to control, significantly higher levels (median [interquartile rang]) of blood pressure (145.5[135.0; 155.0] vs 117.5[110.0; 120.0] mm Hg; P<0.001) NFκB expression in the myocardium (1.25[0.87; 1.87] vs 0.89[0.31; 1.15] relative units; P<0.01) and aorta (0.87[0.57;1.63] vs 0.35[0.22; 0.87] relative units; P<0.05). The expression level of miRNA-21 in the myocardium increased in rats with NE (1.48[0.54; 3.73] vs 0.37[0.06; 1.62] relative units; P<0.01) relative to the SO group, but did not change significantly in the aorta (p>0.05). Relative expression levels of microRNA-133 and -203 both in the myocardium and in the aorta of WKY rats with NE were more than 10 times lower compared to those of the SO group (p<0.001). BMI in rats 2 months after NE increased relative to the SO group (p<0.05). Conclusion An experimental decrease in the number of functioning nephrons not only leads to an increase in blood pressure and myocardial mass in WKY rats, but also causes epigenomic changes in the myocardium and blood vessels, in particular, activation of signaling pathways associated with NFκB in the myocardium and aorta of rats. Possibly, it is accompanied by growth in expression of NFκB-associated proliferative, proinflammatory and profibrotic cytokines. It is possible that microRNA-21 is also involved in the formation of fibrosis and myocardial remodeling. However, specific mechanisms of miRNA involvement in the pathogenesis of heart and vascular remodeling require further study.