Abstract 14472: RNA Sequencing Analyses Reveal Eight miRNA/mRNA Pairs that may Mediate the Effects of Cocaine in Cardiovascular Diseases
Introduction: Cocaine abuse is a significant risk factor for cardiovascular (CV) mortality and morbidity. The underlying molecular mechanisms, however, remain elusive. Goal: MicroRNAs (miRNAs) are short, noncoding RNAs that bind to the 3’UTR of the target mRNA and result in translational repression or degradation. This study aimed to identify miRNA/mRNA pathways that may mediate cocaine’s effects in the CV system.
Methods: C57BL/6 mice were injected (ip) daily with cocaine (20mg/kg body weight), cocaine methiodide (CM, a cocaine analogue that does not enter the CNS), or saline for 10 consecutive days. Total RNA was extracted from aortas (n=3) and subjected to HTSeq-based small RNA sequencing and RNA sequencing containing >41,000 probes. Gene expression profiles between treatment groups (fold changes ≥2) were analyzed, and candidate genes were validated by qRT-PCR. Putative pathways were identified by Gene Ontology and KEGG pathway analyses. miRNA’s targets were predicted using miRDB. The relationship between miRNA and mRNA was determined by co-transfection of HEK293 cells using wt and mutant candidate gene 3’UTR.
Results: When compared to saline treatments, 8 miRNAs were downregulated and 4 miRNAs were upregulated by both cocaine and CM, while 48 genes were upregulated and 267 genes were downregulated by both agents. miRNA prediction analysis revealed 2070 genes to be targets of the 4 upregulated miRNAs, and 3205 genes to be targets of the 8 downregulated miRNAs. Superimposition of the datasets from miRNA prediction and RNAseq revealed 71 genes as predicted targets of the 4 upregulated miRNAs showed decreased expression (anti-correlation) by cocaine and CM, whereas 18 genes as predicted targets of the 8 downregulated miRNAs showed increased expression by cocaine and CM. Based on their relevance to cellular physiology and CV diseases, we ranked 8 top miRNA/mRNA pairs and also established causal relationship between miRNAs and their target genes.
Conclusions: Cocaine appears to affect the homeostasis and functional integrity of aortas by affecting the expression of miRNAs and their target genes. Further functional analyses of the candidate pathways may lead to the discovery of novel molecular targets for intervention for cocaine-induced CV toxicities.
Author Disclosures: W. Zhu: None. H. Wang: None. G.C. Sartor: None. M. Bao: None. A.J. Griswold: None. C.R. Wahlestedt: None. C. Dong: None.
- © 2016 by American Heart Association, Inc.