Abstract 2423: Microrna-34 Cluster Regulates Cardiac Fibroblast Differentiation: Significance in Fibrosis of the Ischemia-Reperfused Heart
Background. microRNA (miR)-34 cluster function as direct and conserved p53 target genes that induce cell cycle arrest. Previously, we have demonstrated that perceived hyperoxia or sub-lethal reoxygenation shock in the peri-infarct tissue during ischemia-reperfusion (I/R) of the heart induces differentiation of cardiac fibroblasts (CF) primarily via an oxygen sensitive p53-p21 pathway. Recently, we have also reported CF specific induction of miR-21 in the ischemia-reperfused heart and demonstrated the significance of miR-21 in post-I/R cardiac fibrosis. The role of miR-34 (a/b/c) cluster in p53-p21 mediated fibroblast differentiation in the I/R heart remains unknown.
Hypothesis. The miR 34 cluster is induced in the I/R heart and regulates fibroblast growth arrest and subsequent differentiation, an important element in myocardial fibrosis.
Methods. Primary CF isolated from adult murine hearts was cultured. As reported earlier, the cells were cultured at 5% O2 (in vivo heart pO2 or normoxia) followed by exposure to 20% O2 (hyperoxia). To study the regulation of miRNA in heart following I/R, mice were subjected to I/R by transient ligation of left anterior descending coronary artery for 60 minutes followed by reperfusion. Infarct and non-infarct tissues were collected at 2 and 7 days post-I/R.
Results. Exposure of CF to 20% O2 (hyperoxic insult) resulted in significant (p<0.05; n=4) induction of miR-34a/b/c expression as well increased activity of a miR-34a reporter. In CF, p53 knock-down resulted in complete abrogation of hyperoxia-inducible miR-34 cluster expression suggesting a central role of p53 in such activation. Anti-miR against miR34a/b/c resulted in partial release and reversal of oxygen-induced growth arrest and differentiation of CF suggesting that along with p21, miR 34-cluster is involved in the fine-regulation of hyperoxia-induced CF growth arrest and subsequent differentiation response. Finally, we observed that the expression of mir34a/b/c is significantly (p<0.05; n=4) induced in the hearts subjected to I/R.
Conclusions. The miR34 cluster is oxygen-sensitive and plays a major role in fine-tuning p53-p21 mediated CF differentiation response. Such regulation may be implicated in cardiac fibrosis response in the I/R heart.