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Identifying microRNAs targeting Wnt/β-catenin pathway in end-stage idiopathic pulmonary arterial hypertension

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Abstract

MicroRNAs (miRNAs) play important roles in the pathogenesis of pulmonary arterial hypertension (PAH). However, the pathways targeted by miRNAs in PAH have not been systematically investigated. We aim to identify dysregulated miRNAs for patients with idiopathic PAH (IPAH). miRNA profiling was performed on lung tissue total RNA from eight IPAH patients and eight control subjects. Real-time quantitative RT-PCR (qRT-PCR) was used for validation of miRNA and mRNA expression levels in 14 IPAH patients and 14 control subjects. Pathway enrichment analysis showed that Wnt/β-catenin signaling is among the top PAH-related pathways enriched in target genes of dysregulated miRNAs. We confirmed the significant increased expression levels of five miRNAs (let-7a-5p, miR-26b-5p, miR-27b-3p, miR-199a-3p and miR-656) targeting major PAH-related pathways. Moreover, qRT-PCR validation of Wnt/β-catenin pathway activation indicated multiple genes including receptors (FZD4, FZD5), core molecule (CTNNB1), and downstream targets (CCND1, VEGFA, and AXIN2) were significantly upregulated. The expression level of miR-199b-5p was positively correlated with patients’ hemodynamics (PVR: r = 0.522, p = 0.038) and pulmonary vascular remodeling (muscularization: r = 0.540, p = 0.021). We confirmed overexpression of miR-199b-5p in hypoxic pulmonary arterial endothelial cells that negatively regulates GSK3B expression. In summary, miRNAs influence the pathogenesis of PAH by regulating major PAH-related pathways including Wnt/β-catenin in end-stage IPAH.

Key message

  • It is the first miRNA profiling study in lung tissue from end-stage idiopathic PAH.

  • We identified dysregulated miRNAs and major pathways (e.g., Wnt signaling) in IPAH.

  • Levels of miRNA expression were correlated with hemodynamics and pathological changes.

  • We observed aberrant expression of target genes in the Wnt/β-catenin pathway.

  • miRNAs influence the pathogenesis of PAH by regulating major PAH-related pathways.

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Acknowledgments

The authors thank the JHMI Deep Sequencing and Array Facility for performing the work on miRNA arrays. The authors thank Brian Graham for pathology assistance and Alan Berger for helpful discussions. Tissue samples were provided by the Genomics Core and Tissue Core under the Pulmonary Hypertension Breakthrough Initiative (PHBI). Funding for the PHBI is provided by the Cardiovascular Medical Research and Education Fund (CMREF). This work was supported by a Johns Hopkins University Microarray Grant to L.G. from Agilent Technologies, and National Institutes of Health (NIH) grant 1R03HL114937 to L.G., K.C.B. and P.M.H. In addition, LG was supported in part by the Gilead Sciences Research Scholars Program in Pulmonary Arterial Hypertension.

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Authors and Affiliations

  1. Department of Cardiopulmonary Circulation, Shanghai Pulmonary Hospital, Tongji University School of Medicine, Shanghai, China

    Danchen Wu & Zhi-Cheng Jing

  2. Division of Allergy and Clinical Immunology, The Johns Hopkins University School of Medicine, Baltimore, MD, USA

    Danchen Wu, Qun Liu, Kathleen C. Barnes & Li Gao

  3. JHMI Deep Sequencing and Microarray Core, The Johns Hopkins University School of Medicine, Baltimore, MD, USA

    C. Conover Talbot Jr.

  4. State Key Laboratory of Cardiovascular Disease, Fu Wai Hospital, Peking Union Medical College and Chinese Academy of Medical Science, Beijing, China

    Zhi-Cheng Jing

  5. Division of Pulmonary and Critical Care Medicine, The Johns Hopkins University School of Medicine, Baltimore, MD, USA

    Rachel L. Damico & Paul M. Hassoun

  6. Division of Pulmonary and Critical Care Medicine, University of Colorado Health Sciences Center, Denver, CO, USA

    Rubin Tuder

  7. The Johns Hopkins Asthma & Allergy Center, 5501 Hopkins Bayview Circle, Baltimore, MD, 21224, USA

    Li Gao

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  1. Danchen Wu
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Correspondence to Li Gao.

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The authors declare that they have no competing interests.

Authors’ contributions

D.W. performed molecular experiments, statistical analysis, and contributed to the manuscript. C.T. performed statistical analysis of the miRNA array data and contributed to the manuscript. Q.L. performed LNA-ISH. R.L.D. performed the forced expression and inhibition of miRNA experiment. Z.J. provided intellectual contributions to the manuscript regarding the role of estrogen and receptors. R.T. performed histopathological analysis and contributed to the manuscript. K.C.B. and P.M.H. provided intellectual contributions to the design of the experiments and contributed to the manuscript. L.G. conceived the project, performed experiments, data analysis, and contributed to the manuscript. All authors have read and approved the final manuscript.

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Wu, D., Talbot, C.C., Liu, Q. et al. Identifying microRNAs targeting Wnt/β-catenin pathway in end-stage idiopathic pulmonary arterial hypertension. J Mol Med 94, 875–885 (2016). https://doi.org/10.1007/s00109-016-1426-z

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  • DOI: https://doi.org/10.1007/s00109-016-1426-z

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