Systems-level regulation of microRNA networks by miR-130/301 promotes pulmonary hypertension
Thomas Bertero, … , Katherine A. Cottrill, Stephen Y. Chan
Thomas Bertero, … , Katherine A. Cottrill, Stephen Y. Chan
Published June 24, 2014
Citation Information: J Clin Invest. 2014;124(8):3514-3528. https://doi.org/10.1172/JCI74773.
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Research Article Pulmonology

Systems-level regulation of microRNA networks by miR-130/301 promotes pulmonary hypertension

Abstract

Development of the vascular disease pulmonary hypertension (PH) involves disparate molecular pathways that span multiple cell types. MicroRNAs (miRNAs) may coordinately regulate PH progression, but the integrative functions of miRNAs in this process have been challenging to define with conventional approaches. Here, analysis of the molecular network architecture specific to PH predicted that the miR-130/301 family is a master regulator of cellular proliferation in PH via regulation of subordinate miRNA pathways with unexpected connections to one another. In validation of this model, diseased pulmonary vessels and plasma from mammalian models and human PH subjects exhibited upregulation of miR-130/301 expression. Evaluation of pulmonary arterial endothelial cells and smooth muscle cells revealed that miR-130/301 targeted PPARγ with distinct consequences. In endothelial cells, miR-130/301 modulated apelin-miR-424/503-FGF2 signaling, while in smooth muscle cells, miR-130/301 modulated STAT3-miR-204 signaling to promote PH-associated phenotypes. In murine models, induction of miR-130/301 promoted pathogenic PH-associated effects, while miR-130/301 inhibition prevented PH pathogenesis. Together, these results provide insight into the systems-level regulation of miRNA-disease gene networks in PH with broad implications for miRNA-based therapeutics in this disease. Furthermore, these findings provide critical validation for the evolving application of network theory to the discovery of the miRNA-based origins of PH and other diseases.

Authors

Thomas Bertero, Yu Lu, Sofia Annis, Andrew Hale, Balkrishen Bhat, Rajan Saggar, Rajeev Saggar, W. Dean Wallace, David J. Ross, Sara O. Vargas, Brian B. Graham, Rahul Kumar, Stephen M. Black, Sohrab Fratz, Jeffrey R. Fineman, James D. West, Kathleen J. Haley, Aaron B. Waxman, B. Nelson Chau, Katherine A. Cottrill, Stephen Y. Chan

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Figure 5

The miR-130/301 family controls PAEC proliferation via the apelin-miR-424/503-FGF2 regulatory axis as well as context-dependent apoptotic signaling.

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The miR-130/301 family controls PAEC proliferation via the apelin-miR-42...
In normoxia (21% O2) or hypoxia (0.2% O2, 24 hours), either forced expression of miR-130a mimic versus control (NC) or inhibition of miR-130a (anti-miR-130a) versus inhibition of the entire miR-130/301 family (tiny-LNA-130) versus control (NC) was performed in cultured PAECs (A–D). Immunoblotting (A), gel densitometry (B), and RT-qPCR (C and D) revealed that forced miR-130a expression decreased PPARγ, a direct target of miR-130/301 (Supplemental Figure 4); decreased apelin (A and B), miR-424 (C), and miR-503 (D); and increased FGF2 (A and B). Inhibition of the entire miR-130/301 family reversed these downstream alterations to a greater extent than miR-130a inhibition alone (A–D), demonstrating the importance of the coordinated actions of this miRNA family. (E) As assessed by BrdU incorporation, PAEC proliferation was augmented by miR-130a mimics but decreased during miR-424 or miR-503 expression. Consistent with the direct dependence of miR-130a on miR-424 and miR-503 in PAECs, miR-130a–induced proliferation was reversed when miR-424, miR-503, or both together were expressed. (F) In serum-starved but not serum-replete PAECs, forced expression of miR-130a increased apoptotic caspase 3/7 activity, while miR-130/301 inhibition partially protected cells from caspase 3/7 induction. (G) In PAECs, the miR-130/301 family controls cellular proliferation as well as context-dependent apoptotic signaling. In C and D, for each miRNA, mean expression in control groups (miR-NC or anti-miR-NC) was assigned a fold change of 1, to which relevant samples were compared. Data are expressed as mean ± SD (*P < 0.05; **P < 0.01).