Gene Expression in Experimental Aortic Coarctation and Repair: Candidate Genes for Therapeutic Intervention?

Abstract

Coarctation of the aorta (CoA) is a constriction of the proximal descending thoracic aorta and is one of the most common congenital cardiovascular defects. Treatments for CoA improve life expectancy, but morbidity persists, particularly due to the development of chronic hypertension (HTN). Identifying the mechanisms of morbidity is difficult in humans due to confounding variables such as age at repair, follow-up duration, coarctation severity and concurrent anomalies. We previously developed an experimental model that replicates aortic pathology in humans with CoA without these confounding variables, and mimics correction at various times using dissolvable suture. Here we present the most comprehensive description of differentially expressed genes (DEGs) to date from the pathology of CoA, which were obtained using this model. Aortic samples (n=4/group) from the ascending aorta that experiences elevated blood pressure (BP) from induction of CoA, and restoration of normal BP after its correction, were analyzed by gene expression microarray, and enriched genes were converted to human orthologues. 51 DEGs with >6 fold-change (FC) were used to determine enriched Gene Ontology terms, altered pathways, and association with National Library of Medicine Medical Subject Headers (MeSH) IDs for HTN, cardiovascular disease (CVD) and CoA. The results generated 18 pathways, 4 of which (cell cycle, immune system, hemostasis and metabolism) were shared with MeSH ID's for HTN and CVD, and individual genes were associated with the CoA MeSH ID. A thorough literature search further uncovered association with contractile, cytoskeletal and regulatory proteins related to excitation-contraction coupling and metabolism that may explain the structural and functional changes observed in our experimental model, and ultimately help to unravel the mechanisms responsible for persistent morbidity after treatment for CoA.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Fig 1. Images of morphological similarity between untreated and corrected CoA in humans (top row) and our rabbit model (bottom row).

Arrows show correction sites. Human images are adapted from related studies discussed in detail in LaDisa et al.—Congenit Heart Dis. 2011 Sep; 6(5): 432–43 and LaDisa and Figueroa et al—J. Biomech. Eng. 2011 Sep;133(9):091008.

Fig 2. Normalized intensity distributions from kernel density plots indicate the sample with a bimodal distribution (Corrected rabbit #18) is an outlier.

This sample was therefore excluded from the analysis indicated by the remaining figures and tables.

Fig 3

(Top) Probes with >2 fold-change and a p-value 8 fold changes. Human genes corresponding to expressed probes on the rabbit chip were determined using orthologue analysis in Better Bunny (Craig et al.—BMC Bioinformatics. 2012 May 8;13:84). Probes without an orthologue human gene and redundant differentially expressed genes (DEGs) were omitted. (Bottom) Unique human orthologue DEGs with >6 FC (indicated within the boxes below) were further studied through GO term, function and pathway analysis, and extensive literature review.

Fig 4. Venn Diagrams of DEGs with >6 FC.

Probes common to the CoA vs Control and Corrected vs Control comparisons could help explain persistent morbidity after restoring BP, as could highly expressed probes in the CoA vs Corrected comparison that are not found in CoA vs Control comparison.

Fig 5. Gene Ontology (GO) terms for DEGs with >6 FC are shown by biological process (top), molecular function (middle), and cellular component (bottom) domains.

Comparison of DEGs was made between groups of samples in three ways: (1) CoA vs Control, (2) Corrected vs Control, and (3) CoA vs Corrected. DEGs common to CoA vs Control and Corrected vs Control are particularly interesting as they could help explain sources of morbidity persisting after restoring BP, as could probes for highly expressed genes in CoA vs Corrected that are not present in the CoA vs Control comparison. These DEGs of potential interesting regions are indicated by black bars, referring back to overlapping DEGs of interest in Fig 4.

Fig 6. Verification of expression by qRT-PCR for two of the genes with the most pronounced decrease in expression by microarray analysis, ITGA4 and UCP1.

* = significantly different (P

Fig 7. Relationship obtained from IPA for the seven DEGs that may collectively account for the hemodynamic, histological, immunohistochemical and myographic changes observed in our rabbit CoA model to date.

Fig 8. Relationship between DEGs within the metabolism pathway of IPA.