Then the role of miR-204 and Tgfbr2 in the anti-PAH effect of 1,25(OH)<sub>2</sub>D<sub>3</sub> was further explored by modulating the expression of the two genes.
Both PAH and CAD are associated with sustained inflammation and smooth muscle cell proliferation/apoptosis imbalance and we demonstrated in PAH that this phenotype is, in part, because of the miR-223/DNA damage/Poly[ADP-ribose] polymerase 1/miR-204 axis activation and subsequent bromodomain protein 4 (BRD4) overexpression.
Using in vitro gain- and loss-of-function approaches, we mechanistically demonstrate that miR-204 diminution promotes RUNX2 up-regulation and that sustained RUNX2 expression activates hypoxia-inducible factor-1α, leading to aberrant proliferation, resistance to apoptosis, and subsequent transdifferentiation of PAH-PASMCs into osteoblast-like cells.
We also showed that PARP-1 activation accounts for miR-204 downregulation (quantitative reverse transcription polymerase chain reaction) and the subsequent activation of the transcription factors nuclear factor of activated T cells and hypoxia-inducible factor 1-α in PAH-PASMCs, previously shown to be critical for PAH in several models.
Moreover, through the down-regulation of miR-204, STAT3 enhances a positive feedback loop sustaining its own activation, showing that miRNA regulation is critical in PAH.
This study uncovers a new regulatory pathway involving miR-204 that is critical to the etiology of PAH and indicates that reestablishing miR-204 expression should be explored as a potential new therapy for this disease.