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  • microRNA overexpression in slow transit constipation leads to reduced NaV1.5 current and altered smooth muscle contractility

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Neurogastroenterology
Original research
microRNA overexpression in slow transit constipation leads to reduced NaV1.5 current and altered smooth muscle contractility
  1. Amelia Mazzone1,
  2. Peter R Strege1,2,
  3. Simon J Gibbons1,2,
  4. Constanza Alcaino1,
  5. Vikram Joshi1,
  6. Andrew J Haak2,
  7. Daniel J Tschumperlin2,
  8. Cheryl E Bernard1,
  9. Robert R Cima3,
  10. David W Larson3,
  11. Heidi K Chua3,
  12. Rondell P Graham4,
  13. Mona El Refaey5,6,
  14. Peter J Mohler5,
  15. Yujiro Hayashi1,
  16. Tamas Ordog1,2,
  17. Stefan Calder7,
  18. Peng Du7,
  19. Gianrico Farrugia1,2,
  20. Arthur Beyder1,2
  1. 1 Enteric NeuroScience Program (ENSP), Division of Gastroenterology and Hepatology, Department of Medicine, Mayo Clinic, Rochester, Minnesota, USA
  2. 2 Department of Physiology and Biomedical Engineering, Mayo Clinic, Rochester, Minnesota, USA
  3. 3 Department of Colon and Rectal Surgery, Mayo Clinic, Rochester, Minnesota, USA
  4. 4 Department of Pathology, Mayo Clinic, Rochester, Minnesota, USA
  5. 5 Departments of Physiology and Cell Biology, The Dorothy M. Davis Heart and Lung Research Institute, The Ohio State University Wexner Medical Center, Columbus, Ohio, USA
  6. 6 Department of Internal Medicine, The Dorothy M. Davis Heart and Lung Research Institute, The Ohio State University Wexner Medical Center, Columbus, Ohio, USA
  7. 7 Auckland Bioengineering Institute, University of Auckland, Auckland, New Zealand
  1. Correspondence to Dr Gianrico Farrugia, Enteric NeuroScience Program, Division of Gastroenterology and Hepatology, Department of Medicine, Mayo Clinic, Rochester, MN 55905, USA; farrugia.gianrico{at}mayo.edu; Dr Arthur Beyder, Enteric NeuroScience Program, Division of Gastroenterology and Hepatology, Department of Medicine, Mayo Clinic, Rochester, MN, 55905; beyder.arthur{at}mayo.edu

Abstract

Objective This study was designed to evaluate the roles of microRNAs (miRNAs) in slow transit constipation (STC).

Design All human tissue samples were from the muscularis externa of the colon. Expression of 372 miRNAs was examined in a discovery cohort of four patients with STC versus three age/sex-matched controls by a quantitative PCR array. Upregulated miRNAs were examined by quantitative reverse transcription qPCR (RT-qPCR) in a validation cohort of seven patients with STC and age/sex-matched controls. The effect of a highly differentially expressed miRNA on a custom human smooth muscle cell line was examined in vitro by RT-qPCR, electrophysiology, traction force microscopy, and ex vivo by lentiviral transduction in rat muscularis externa organotypic cultures.

Results The expression of 13 miRNAs was increased in STC samples. Of those miRNAs, four were predicted to target SCN5A, the gene that encodes the Na+ channel NaV1.5. The expression of SCN5A mRNA was decreased in STC samples. Let-7f significantly decreased Na+ current density in vitro in human smooth muscle cells. In rat muscularis externa organotypic cultures, overexpression of let-7f resulted in reduced frequency and amplitude of contraction.

Conclusions A small group of miRNAs is upregulated in STC, and many of these miRNAs target the SCN5A-encoded Na+ channel NaV1.5. Within this set, a novel NaV1.5 regulator, let-7f, resulted in decreased NaV1.5 expression, current density and reduced motility of GI smooth muscle. These results suggest NaV1.5 and miRNAs as novel diagnostic and potential therapeutic targets in STC.

  • motility disorders
  • intestinal motility
  • intestinal ion transport
  • genetics
  • constipation
http://creativecommons.org/licenses/by-nc/4.0/

This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited, appropriate credit is given, any changes made indicated, and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/.

https://doi.org/10.1136/gutjnl-2019-318747

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    Footnotes

    • GF and AB are joint senior authors.

    • Twitter @beyderlab

    • Contributors AM designed and performed experiments, analysed data and wrote the manuscript. PRS, VJ, YH, MER, CA and AJH performed experiments and analysed data; TO, PJM, SC, DT, RG, SJG and PD designed experiments and analysed data; CEB coordinated identification and retrieval of surgical tissue; RRC, DWL and HKC performed surgeries and provided tissues; AB and GF designed the research, analysed data and wrote the manuscript. All authors critically reviewed and approved the manuscript.

    • Funding This study was funded by National Institute of Diabetes and Digestive and Kidney Diseases NIH R01 (DK52766) to GF and by NIH K08 (DK106456), American Gastroenterological Association Research Scholar Award (AGA RSA) to AB and The Rutherford Discovery Fellowship by The Rutherford Foundation, Royal of Society of New Zealand to PD.

    • Competing interests None declared.

    • Patient consent for publication Not required.

    • Provenance and peer review Not commissioned; externally peer reviewed.

    • Data availability statement All data relevant to the study are included in the article or uploaded as supplementary information.