miR-127-3p is an epigenetic activator of myofibroblast senescence situated within the miRNA enriched Dlk1-Dio3 imprinted domain on mouse chromosome 12 [mRNA array]

During wound healing, fibroblasts differentiate into non-proliferative contractile myofibroblasts, contribute to skin repair, and eventually undergo apoptosis or become senescent. MicroRNAs (miR) are posttranscriptional regulators of gene expression networks that control cell fate and survival and may also regulate senescence. Here we determined regulated miRs in myofibroblasts isolated from wounds and analyzed their role in senescent myofibroblast formation. Transcriptome profiling showed that a 200 kbp region of the Dlk1-Dio3 imprinted domain on mouse chromosome 12 encodes for most of the upregulated miRs in the entire genome of mouse myofibroblasts. Among those, miR-127-3p induced a myofibroblast-like phenotype associated with a block in proliferation. Molecular analysis revealed that miR-127-3p induced a prolonged cell-cycle arrest with unique molecular features of senescence, including the activation of the senescence-associated ß-galactosidase, increase in p21 levels, inhibition of lamin B1, proliferation factors, and the production of senescence-associated inflammatory and extracellular matrix -remodeling components. Hence, miR-127-3p emerges as an epigenetic activator regulating the transition from repair to remodeling during skin wound healing, but may also induce age-related defects, pathological scarring and fibrosis, all linked to myofibroblast senescence. Dermal cells were isolated from skin of C57BL/6 newborn mice following dispase and collagenase type 1 digest as described (Etich et al., 2013) and cultured in DMEM (Gibco/ Thermo Fisher Scientific, Waltham, USA) with 10% FCS and 1% P/S. For transfection experiments 1x105 cells were cultured in a 12-well-plate and transfected with control or miR-127-3p mimic oligonucleotides. Phenol-chloroform extracted non-degraded RNA (100 ng) isolated from control or miR-127-3p transfected fibroblasts was amplified, labeled using the Low input Quick Amp Labeling kit and hybridized to a SurePrint G3 Mouse GE 8x60K Microarray (Agilent, Santa Clara, USA). After scanning and processing data were analyzed using Genespring14.9.1 Multi-Omic software and FunRich tool (Pathan et al., 2015).