Differential Expression of microRNAs in Severely Calcified Carotid Plaques

Background: We investigated whether microRNA (miRNA) alteration is related to the presence of calcification in carotid plaques. Methods: We classified 10 plaques from carotid endarterectomy patients into high- and low-calcified plaques based on Agatston calcium scores. A microarray analysis for miRNA profiles was performed, with validation by a miRNA quantitative real-time polymerase chain reaction (qRT-PCR). Results: The miRNA microarray identified 697 probes; 657 of them were downregulated. We selected the genes that satisfied total gene signal (TGS) >50, |Log2 ratio | > 1 >= 1 and of the following: (1) false discovery rate (FDR) <.05 in the comparison of mean values of logarithmic transformed signals between the groups; (2) .05 <= FDR < .1 and showing either high or median for context score+ in miRSearch among the 72 carefully selected genes related to angiogenesis or calcification; and (3) FDR < .1 in the comparison of 10 individual sets of high- and low-calcified plaques. The expression of miRNA validated by qRT-PCR revealed a significant downregulation of hsa-miR-4530, hsa-miR133b, and hsa-miR-1-3p. A Spearman's rank correlation analysis revealed that the logarithmic TGSs for the microarray of hsa-miR-4530 and hsa-miR-133b were significantly inversely correlated with the carotid plaques' calcium scores, and the delta Cq values for the qRT-PCR showed a direct association. Conclusions: In high-calcified carotid plaques, a specific profile for miRNA may be identified, and the expressions of hsa-miR-4530 and hsa-miR-133b had inverse correlations with the calcium score in the plaques, suggesting that miRNAs may play a modulating role in calcified plaques and plaque stability.