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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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 8

The miR-130/301 family is necessary to induce PH in a hypoxic mouse model of disease.

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The miR-130/301 family is necessary to induce PH in a hypoxic mouse mode...
A protocol was performed to determine whether a “shortmer” inhibitor of miR-130/301 (short-130) reverses PH (hypoxia + SU5416) in mice. See Supplemental Figure 23. (A) While miR-130/301 progressively increased with exposure time to hypoxia + SU5416, short-130 maintained baseline miRNA expression (normoxia + SU5416). (B) RVSP progressively increased as mice were exposed to 2 weeks (Ctrl 2 weeks) and 4 weeks of hypoxia + SU5416 (PBS). Increased RVSP was abrogated by short-130 but not short-NC. See Supplemental Figure 26A for quantification of (RV/LV+S) mass ratio. (C) During hypoxia + SU5416, short-130 rescued miR-204, miR-322 (mouse homolog of human miR-424), and miR-503 expression in whole lung. (D) IHC staining of pulmonary vessels less than 100 μm diameter was performed to quantify PPARγ, PCNA, phosphorylated STAT3 (P-STAT3), and α-SMA. Ten vessels were quantified for each mouse (bar graphs). Inhibition of the miR-130/301 family after PH development preserved PPARγ expression (top row); decreased PCNA-positive (second row) and P-STAT3–positive (third row) cells (quantified over 10 vessels); and decreased medial thickness (bottom row) consistent with blunted vascular remodeling. See Supplemental Figure 26B for arteriolar density, Supplemental Figure 26C for arteriolar muscularization, and Supplemental Figure 26, D and E, for immunoblotting. In A and C, for each miRNA, normoxic levels were assigned a fold change of 1, to which relevant samples were compared. Data involving animal tissue (A–C) are expressed as mean ± SEM, while IHC quantification (D) is expressed as mean ± SD (*P < 0.05; **P < 0.01). Scale bar: 50 μm.