Longitudinal Analysis of Gene Expression in Porcine Skeletal Muscle After Post-Injection Local Injury
The purpose of this study is to describe the time course of gene expression in a skeletal muscle local injury induced by an intramuscular (IM) injection, and to compare the dynamics of gene expression with pathological events.
Materials and Methods
Ten piglets received 4 IM injections of propylene glycol in the longissimus dorsi muscles 6 h, 2, 7, and 21 days before euthanasia, where control and injected muscle sites were sampled for RNA isolation and microscopic examination. The hybridization of nylon cDNA microarrays was carried out with radioactive probes obtained from the muscle RNA.
153 genes were found under- or over-expressed at least once among the investigated time-conditions. The eight most discriminant genes were also identified: Two genes (GTP-binding protein RAD and Ankyrin repeat domain protein) were over-expressed at 6 h and six genes between 2 and 21 days (Osteonectin, Fibronectin, Matrix metalloproteinase-2, Collagen alpha 1(I) chain, Collagen alpha 2(I) chain, and Thymosin beta-4). Necrosis, inflammation and regeneration were observed through both the dynamics of gene expression profiles and through the microscopic examinations.
Our data demonstrate that several pathways are involved in post-injection muscle injury, and that necrosis, inflammation and regeneration are not sequential but occur in parallel.
Key wordsgene expression profiling injury intramuscular injection local tolerance pathophysiology
The muscle specific cDNA library used in this work was kindly provided by Christian Bendixen. The authors thank Rémi Houlgatte (TAGC, Marseille, France) for critical discussions about/on Nylon microarray methodology and data analysis. We thank Jean-Pierre Gau, Nadine Gautier, Francis Benne, and Janine Rallières for their technical assistance. We wish to acknowledge support from the CRGS platform of the Toulouse Midi-Pyrenees Genopole (Cécile Donnadieu-Tonon) (http://genopole-toulouse.prd.fr/), where the nylon membranes were produced. The database with BASE software for MIAME submission was developed by Christelle Dantec and managed by a computer group (SIGENAE, Système d’information du projet d’analyse des génomes des animaux d’élevage, http://www.sigenae.org). We thank the work group for statistical analysis of microarray data (Philippe Besse, Alain Baccini, Sébastien Déjean, http://www.lsp.ups-tlse.fr/Programmes/Biopuces/) for their advice in the RF analysis and to Christèle Robert-Granié for helpful discussions on the statistical methods.
- 14.A. Brazma, P. Hingamp, J. Quackenbush, G. Sherlock, P. Spellman, C. Stoeckert, J. Aach, W. Ansorge, C. A. Ball, H. C. Causton, T. Gaasterland, P. Glenisson, F. C. Holstege, I. F. Kim, V. Markowitz, J. C. Matese, H. Parkinson, A. Robinson, U. Sarkans, S. Schulze-Kremer, J. Stewart, R. Taylor, J. Vilo, and M. Vingron. Minimum information about a microarray experiment (MIAME)-toward standards for microarray data. Nat. Genet. 29:365–371 (2001).PubMedCrossRefGoogle Scholar
- 19.L. H. Saal, C. Troein, J. Vallon-Christersson, S. Gruvberger, A. Borg, and C. Peterson. BioArray Software Environment (BASE): a platform for comprehensive management and analysis of microarray data. Genome. Biol. 3:SOFTWARE0003 (2002).Google Scholar
- 22.K. Sudre, C. Leroux, G. Pietu, I. Cassar-Malek, E. Petit, A. Listrat, C. Auffray, B. Picard, P. Martin, and J. F. Hocquette. Transcriptome analysis of two bovine muscles during ontogenesis. J. Biochem. (Tokyo) 133:745–756 (2003).Google Scholar
- 27.P. J. Ferre, D. Concordet, V. Laroute, G. P. Chanoit, J. P. Ferre, M. Manesse, and H. P. Lefebvre. Comparison of ultrasonography and pharmacokinetic analysis of creatine kinase release for quantitative assessment of postinjection muscle damage in sheep. Am. J. Vet. Res. 62:1698–1705 (2001).PubMedCrossRefGoogle Scholar
- 34.C. Strup-Perrot, D. Mathe, C. Linard, D. Violot, F. Milliat, A. Francois, J. Bourhis, and M. C. Vozenin-Brotons. Global gene expression profiles reveal an increase in mRNA levels of collagens, MMPs, and TIMPs in late radiation enteritis. Am. J. Physiol. Gastrointest. Liver Physiol. 287:G875–G885 (2004).PubMedCrossRefGoogle Scholar