Abstract
Clenbuterol is currently being used as part of a clinical trial into a novel therapeutic approach for the treatment of end-stage heart failure. The purpose of this study was to determine the global pattern of myocardial gene expression in response to clenbuterol and to identify novel targets and pathways involved. Rats were treated with clenbuterol (n = 6) or saline (n = 6) for periods of 1, 3, 9, or 28 days. Rats treated for 28 days were also subject to continuous electrocardiogram analysis using implantable telemetry. RNA was extracted from rats at days 1 and 28 and used from microarray analysis, and further samples from rats at days 1, 3, 9, and 28 were used for analysis by real-time polymerase chain reaction. Clenbuterol treatment induced rapid development of cardiac hypertrophy with increased muscle mass at day 1 and elevated heart rate and QT interval throughout the 28-day period. Microarray analysis revealed a marked but largely transitory change in gene expression with 1,423 genes up-regulated and 964 genes down-regulated at day 1. Up-regulated genes revealed an unexpected association with angiogenesis and integrin-mediated cell adhesion and signaling. Moreover, direct treatment of endothelial cells cultured in vitro resulted in increased cell proliferation and tube formation. Our data show that clenbuterol treatment is associated with rapid cardiac hypertrophy and identify angiogenesis and integrin signaling as novel pathways of clenbuterol action. The data have implications both for our understanding of the physiologic hypertrophy induced by clenbuterol and for treatment of heart failure.
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This work was supported by the Magdi Yacoub Institute.
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Below is the link to the electronic supplementary material.Lara-Pezzi et al., A gene expression profile of the myocardial response to clenbuterol. Supplementary data information.
Supplementary Tables 1–4
Triplicate samples of clenbuterol treated and saline treated controls from each of the days 1 and 28 day groups were analyzed using Affymetrix DNA oligonucleotide RAE230 2.0 microchips (Affymetrix, Santa Clara, CA, USA) and data analyzed using the Genespring software suite (Agilent, Santa Clara, CA, USA) as described. Lists of genes altered in response to 1 or 28 days of clenbuterol treatment are listed; individual Excel gene lists of genes up-regulated at 1 day (Up clen 1d), down-regulated at 1 day (Down clen 1 d), or up- or down-regulated at 28 days (Up clen 28 d and Down clen 28 d respectively). (XLS 381 KB)
Supplementary Tables 5–8
Triplicate samples of clenbuterol-treated and saline-treated controls following either 1 or 28 days of treatment were analyzed using Affymetrix DNA oligonucleotide as above. Gene lists were created in which all genes showed raw expression values >100 in two out of four conditions (1 and 28 days, ±clenbuterol), flags were present or marginal in at least three samples, and gene expression showed a fold change of >1.4-fold. The resulting lists were compared to pre-existing Gene Ontology lists using Genespring Analysis (Agilent, Santa Clara, CA, USA). Resulting data are shown in Supplementary Tables 5–8 for sample sets for genes up-regulated or down-regulated at day 1 (Up clen d 1, Down clen day 1, respectively) and similarly for day 28 (Up clen 28 and Down clen 28 d) and listed under Biological process, Cellular Compartment and Molecular Function. (XLS 11.5 KB)
Supplementary Tables 9–12
Triplicate samples of clenbuterol treated and saline treated controls were analyzed using Affymetrix DNA oligonucleotide as above, and the resulting gene lists used (as defined under Supplementary Tables 5–8) analyzed using Ingenuity Pathways Analysis (Ingenuity, www.ingenuity.com). (XLS 311 KB)
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Lara-Pezzi, E., Terracciano, C.M.N., Soppa, G.K.R. et al. A Gene Expression Profile of the Myocardial Response to Clenbuterol. J. of Cardiovasc. Trans. Res. 2, 191–197 (2009). https://doi.org/10.1007/s12265-009-9097-6
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DOI: https://doi.org/10.1007/s12265-009-9097-6