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Biologia

, Volume 74, Issue 12, pp 1687–1698 | Cite as

Validation of reference genes in human epicardial adipose tissue and left ventricular myocardium in heart failure

  • Jana Mlynarova
  • Andrea GazovaEmail author
  • Peter Musil
  • Andrea Raganová
  • Ondrej Sprusansky
  • Eva Goncalvesova
  • Michal Hulman
  • John J. Leddy
  • Jan Kyselovic
Original Article
  • 19 Downloads

Abstract

The characterization of tissue-specific gene expression in failing hearts requires the identification of appropriately validated reference genes. This study sought to validate which commonly used reference genes were the most suitable for qRT-PCR data normalization in epicardial adipose tissue and left ventricular myocardium in patients with end-stage heart failure. The mRNA expression of 10 candidate reference genes was analyzed by qRT-PCR. The stability of their expression was evaluated by our novel scoring system and compared with known algorithms used to identify stable reference genes (Normfinder and Bestkeeper). Using these three methods, we selected the Ribosomal protein L13A (RPL13A) as the best reference gene when studying either epicardial adipose tissue or left ventricular myocardium in failing hearts. In contrast, the β2-microglobulin (B2M) gene was identified as a more suitable reference gene in studies comparing gene expression in epicardial adipose tissue to left ventricular myocardium.

Keywords

qRT-PCR normalization Ribosomal protein L13A β2-microglobulin 

Abbreviations

Yrs

years

EAT

epicardial adipose tissue

B2M

β2-microglobulin

GAPDH

Glyceraldehyde-3-phosphate dehydrogenase

GUSB

β-glucuronidase

ACTB

β-actin

HPRT1

Hypoxanthine phosphoribosyltransferase 1

LRP10

Low-density lipoprotein receptor-related protein

PGK1

Phosphoglycerate kinase 1

RPII

RNA polymerase II

RPL13A

Ribosomal protein L13A

TFRC

Transferrin receptor

LV

left ventricular myocardium

Notes

Acknowledgments

This publication is the result of the project implementation: Biomakro ITMS: 26240120027, VEGA 1/0905/14, VEGA 1/0949/15, APVV-14-0416.

Special thanks to colleagues Peter Krenek, Gabriel Dóka and Lenka Pivackova.

Compliance with ethical standards

Conflict of interest

The authors declare no conflict of interest.

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Copyright information

© Institute of Molecular Biology, Slovak Academy of Sciences 2019 2019

Authors and Affiliations

  1. 1.V. Department of Internal MedicineFaculty of Medicine, Comenius UniversityBratislavaSlovakia
  2. 2.Institute of Pharmacology and Clinical PharmacologyFaculty of Medicine, Comenius UniversityBratislavaSlovakia
  3. 3.Department of Pharmacology and Toxicology, Faculty of PharmacyComenius UniversityBratislavaSlovakia
  4. 4.National Institute for Cardiovascular DiseasesBratislavaSlovakia
  5. 5.Department of Cellular and Molecular Medicine, Faculty of MedicineUniversity of OttawaOttawaCanada

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