UNBIASED ASSESSMENT OF CIRCULATING MICRORNAS PROVIDES AN INDEPENDENT LINK BETWEEN THE CHOLESTEROL-REGULATING MIR-382-5P AND CORONARY ARTERY DISEASE IN HUMANS

Abstract

BackgroundMicroRNAs are short non-protein coding RNA molecules with pervasive regulatory roles in cardiovascular biology. miR-382-5p has recently been shown to contribute to the regulation of cholesterol homeostasis and inflammatory response in mouse models of atherosclerosis; however, the relevance of this miRNA to human cardiovascular disease is not yet known. Because cellular miRNAs can be released into circulation in a stable extracellular format, the assessment of plasma miRNA levels may provide unique insight into underlying disease activity in patients with cardiovascular disease.ObjectiveTo identify circulating extracellular miRNAs that are differentially altered between coronary artery disease (CAD) patients and healthy control subjects.MethodsTotal RNA was extracted from 200 uL of citrate plasma derived from peripheral blood of 5 CAD patients (mean age 55 ± 6 yr, 100 % female) and 6 healthy control subjects (mean age 50 ± 4 yr, 100% female). A total of 372 different miRNAs were quantified using an unbiased high density PCR array strategy (miScript, Qiagen), and data were normalized with a mean-centering restricted approach.ResultsThe levels of 21 miRNAs were altered between CAD patients and healthy subjects (P<0.05), including 19 miRNAs that were increased in CAD patients (from 1.3 - 2.9 fold) and 2 miRNAs that were decreased (from 1.6-1.7 fold). Unsupervised hierarchical clustering with this subset of altered miRNAs showed clear separation of CAD patients and controls. miR-382-5p was the most significantly altered miRNA (1.8 fold up in CAD, P=0.0004), and exhibited an area under the corresponding receiver operator characteristic curve of 1.0 (P = 0.006), indicative of a potential biomarker with exceptional sensitivity and specificity.ConclusionsAn unbiased assessment of circulating miRNAs identified elevated levels of the cholesterol-regulating miR-382-5p in CAD patients. To the best of our knowledge, this is the first evidence directly linking miR-382-5p to cardiovascular disease in humans. BackgroundMicroRNAs are short non-protein coding RNA molecules with pervasive regulatory roles in cardiovascular biology. miR-382-5p has recently been shown to contribute to the regulation of cholesterol homeostasis and inflammatory response in mouse models of atherosclerosis; however, the relevance of this miRNA to human cardiovascular disease is not yet known. Because cellular miRNAs can be released into circulation in a stable extracellular format, the assessment of plasma miRNA levels may provide unique insight into underlying disease activity in patients with cardiovascular disease. MicroRNAs are short non-protein coding RNA molecules with pervasive regulatory roles in cardiovascular biology. miR-382-5p has recently been shown to contribute to the regulation of cholesterol homeostasis and inflammatory response in mouse models of atherosclerosis; however, the relevance of this miRNA to human cardiovascular disease is not yet known. Because cellular miRNAs can be released into circulation in a stable extracellular format, the assessment of plasma miRNA levels may provide unique insight into underlying disease activity in patients with cardiovascular disease. ObjectiveTo identify circulating extracellular miRNAs that are differentially altered between coronary artery disease (CAD) patients and healthy control subjects. To identify circulating extracellular miRNAs that are differentially altered between coronary artery disease (CAD) patients and healthy control subjects. MethodsTotal RNA was extracted from 200 uL of citrate plasma derived from peripheral blood of 5 CAD patients (mean age 55 ± 6 yr, 100 % female) and 6 healthy control subjects (mean age 50 ± 4 yr, 100% female). A total of 372 different miRNAs were quantified using an unbiased high density PCR array strategy (miScript, Qiagen), and data were normalized with a mean-centering restricted approach. Total RNA was extracted from 200 uL of citrate plasma derived from peripheral blood of 5 CAD patients (mean age 55 ± 6 yr, 100 % female) and 6 healthy control subjects (mean age 50 ± 4 yr, 100% female). A total of 372 different miRNAs were quantified using an unbiased high density PCR array strategy (miScript, Qiagen), and data were normalized with a mean-centering restricted approach. ResultsThe levels of 21 miRNAs were altered between CAD patients and healthy subjects (P<0.05), including 19 miRNAs that were increased in CAD patients (from 1.3 - 2.9 fold) and 2 miRNAs that were decreased (from 1.6-1.7 fold). Unsupervised hierarchical clustering with this subset of altered miRNAs showed clear separation of CAD patients and controls. miR-382-5p was the most significantly altered miRNA (1.8 fold up in CAD, P=0.0004), and exhibited an area under the corresponding receiver operator characteristic curve of 1.0 (P = 0.006), indicative of a potential biomarker with exceptional sensitivity and specificity. The levels of 21 miRNAs were altered between CAD patients and healthy subjects (P<0.05), including 19 miRNAs that were increased in CAD patients (from 1.3 - 2.9 fold) and 2 miRNAs that were decreased (from 1.6-1.7 fold). Unsupervised hierarchical clustering with this subset of altered miRNAs showed clear separation of CAD patients and controls. miR-382-5p was the most significantly altered miRNA (1.8 fold up in CAD, P=0.0004), and exhibited an area under the corresponding receiver operator characteristic curve of 1.0 (P = 0.006), indicative of a potential biomarker with exceptional sensitivity and specificity. ConclusionsAn unbiased assessment of circulating miRNAs identified elevated levels of the cholesterol-regulating miR-382-5p in CAD patients. To the best of our knowledge, this is the first evidence directly linking miR-382-5p to cardiovascular disease in humans. An unbiased assessment of circulating miRNAs identified elevated levels of the cholesterol-regulating miR-382-5p in CAD patients. To the best of our knowledge, this is the first evidence directly linking miR-382-5p to cardiovascular disease in humans.