MicroRNA-140-5p and SMURF1 regulate pulmonary arterial hypertension
Alexander M.K. Rothman ... David J. Rowlands, Allan Lawrie
Alexander M.K. Rothman ... David J. Rowlands, Allan Lawrie
Published July 1, 2016
Citation Information: J Clin Invest. 2016;126(7):2495-2508. doi:10.1172/JCI83361.
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Categories: Research Article Pulmonology

MicroRNA-140-5p and SMURF1 regulate pulmonary arterial hypertension

Abstract

Loss of the growth-suppressive effects of bone morphogenetic protein (BMP) signaling has been demonstrated to promote pulmonary arterial endothelial cell dysfunction and induce pulmonary arterial smooth muscle cell (PASMC) proliferation, leading to the development of pulmonary arterial hypertension (PAH). MicroRNAs (miRs) mediate higher order regulation of cellular function through coordinated modulation of mRNA targets; however, miR expression is altered by disease development and drug therapy. Here, we examined treatment-naive patients and experimental models of PAH and identified a reduction in the levels of miR-140-5p. Inhibition of miR-140-5p promoted PASMC proliferation and migration in vitro. In rat models of PAH, nebulized delivery of miR-140-5p mimic prevented the development of PAH and attenuated the progression of established PAH. Network and pathway analysis identified SMAD-specific E3 ubiquitin protein ligase 1 (SMURF1) as a key miR-140-5p target and regulator of BMP signaling. Evaluation of human tissue revealed that SMURF1 is increased in patients with PAH. miR-140-5p mimic or SMURF1 knockdown in PASMCs altered BMP signaling, further supporting these factors as regulators of BMP signaling. Finally, Smurf1 deletion protected mice from PAH, demonstrating a critical role in disease development. Together, these studies identify both miR-140-5p and SMURF1 as key regulators of disease pathology and as potential therapeutic targets for the treatment of PAH.

Authors

Alexander M.K. Rothman, Nadine D. Arnold, Josephine A. Pickworth, James Iremonger, Loredana Ciuclan, Robert M.H. Allen, Sabine Guth-Gundel, Mark Southwood, Nicholas W. Morrell, Matthew Thomas, Sheila E. Francis, David J. Rowlands, Allan Lawrie

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Figure 1

miR-140-5p is reduced in patients with PH and animal models of PAH and modulates PASMC phenotype.

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miR-140-5p is reduced in patients with PH and animal models of PAH and m...
(A) Heat map representing differentially expressed whole blood miR in treatment-naive patients with PH and control groups (fold change > 2, comparison between patients with PH and control groups, n = 4 per group). (B) Effect of whole blood miR on PASMC proliferation in vitro. PASMCs were transfected with miR mimic, inhibitor (Inh), or scramble control (SCR) and proliferation assessed at 72 hours. Inhibition of miR-135a led to cell death, and inhibition of miR-140-5p led to increased proliferation (n = 5, *P < 0.05, 1-way ANOVA with Dunnett’s post-test correction, mean ± SEM). (C and D) PASMC proliferation (C) and migration (D) following miR-140-5p mimic and inhibitor transfection at 72 and 6 hours, respectively. miR-140-5p inhibitor led to increased proliferation and migration in comparison with both SCR and miR-140-5p mimic (C: n = 5, D: n = 4, C and D: *P < 0.05, **P < 0.01, 1-way ANOVA with Tukey’s post-test correction, mean ± SEM, photomicrographs representative of n = 5 (C), and n = 4 (D). Original magnification, ×10 (C); ×20 (D). (E and F) miR-140-5p is reduced during development of PAH in the MCT rat model. RVSP (E) is increased and miR-140-5p (F) decreased at day 28 following injection of 60 mg/kg MCT in the Sprague-Dawley rat (E and F, n = 5–8 per group, *P < 0.05, 2-way ANOVA with Tukey’s post-test correction, mean ± SEM). (G and H) miR-140-5p is reduced during the development of PAH in the SuHx rat model. RVSP is increased and miR-140-5p decreased at week 14 following injection with 20 mg/kg Su5416 and 3 weeks of 10% O2 in the Wistar-Kyoto rat (G and H: n = 8 per group, *P < 0.05, 2-way ANOVA with Tukey’s post-test correction, mean ± SEM).