Abstract 1700: miRNA-dependent Regulation of Smooth Muscle Cell Function During Neointima Formation
Introduction: Micro RNAs (miRNAs) are implicated to regulate mRNA levels of up to 30% of mammalian genes, comprising key regulators for cellular function. Maturation and expression of miRNAs is tightly controlled by the two processing enzymes Dicer and Drosha. Up to now, the role of miRNAs for vascular smooth muscle cell (VSMC) function remains elusive. Thus, the aim of this study was to evaluate the regulation and impact of Dicer and Drosha for smooth muscle cell function during the development of neointimal lesions in vitro and in vivo.
Methods and Results: Following wire-induced injury of the femoral artery of C57BL/6N mice, Dicer or Drosha mRNA expression was analyzed by real-time PCR in the developing neointimal lesions. Expression of Dicer and Drosha was significantly down regulated at day 10 and 21 following injury. Microarray analysis of miRNA expression at these time points revealed a significant downregulation of miRNAs implicated to regulate key molecules of cell cycle progression. Indeed, siRNA-mediated knock down of Dicer or Drosha in human coronary SMC significantly augmented the growth factor-induced expression of CDK4, CDK6, SKP2, accelerated cell cycle progression as determined by FACS analysis and increased VSMC proliferation as determined by total cell count as well as BrdU incorporation (Drosha: 0.0983 ± 0.0194 vs. 0.0527 ± 0.0115; Dicer: 0.0945 ± 0.0172 vs. 0.0527 ± 0.0115; P < 0.005). In contrast, overexpression of Drosha and Dicer significantly reduced VSMC proliferation. Interestingly, knock down of Dicer and Drosha had no effect on migration or apoptosis. Since growth factor stimulation or oxidative stress had no effect, TNF α, IL-1β and IFNγ were identified as potent regulators of Drosha and Dicer expression.
Conclusions: These data indicate that the key miRNA-processing enzymes Dicer and Drosha are down regulated in neointimal lesions, most likely by pro-inflammatory cytokines. Moreover, altered expression of these enzymes and of miRNAs seems to regulate VSMC function during the development of vascular proliferative diseases.