Abstract
Impaired regulation of the transforming growth factor-β (TGFβ) signaling pathway has been linked to thoracic aortic aneurysm (TAA). Previous work has indicated that differential splicing is a common phenomenon, potentially influencing the function of proteins. In the present study we investigated the occurrence of differential splicing in the TGFβ pathway associated with TAA in patients with bicuspid aortic valve (BAV) and tricuspid aortic valve (TAV). Affymetrix human exon arrays were applied to 81 intima/media tissue samples from dilated (n = 51) and nondilated (n = 30) aortas of TAV and BAV patients. To analyze the occurrence of alternative splicing in the TGFβ pathway, multivariate techniques, including principal component analysis and OPLS-DA (orthogonal partial least squares to latent structures discriminant analysis), were applied on all exons (n = 614) of the TGFβ pathway. The scores plot, based on the splice index of individual exons, showed separate clusters of patients with both dilated and nondilated aorta, thereby illustrating the potential importance of alternative splicing in TAA. In total, differential splicing was detected in 187 exons. Furthermore, the pattern of alternative splicing is clearly differs between TAV and BAV patients. Differential splicing was specific for BAV and TAV patients in 40 and 86 exons, respectively, and splicings of 61 exons were shared between the two phenotypes. The occurrence of differential splicing was demonstrated in selected genes by reverse transcription-polymerase chain reaction. In summary, alternative splicing is a common feature of TAA formation. Our results suggest that dilatation in TAV and BAV patients has different alternative splicing fingerprints in the TGFβ pathway.
Highlights
Thoracic aortic aneurysm (TAA) is a pathological widening of the aorta resulting from degeneration of the extracellular matrix and loss of smooth muscle cells in the tunica media
orthogonal projections to latent structures discriminant analysis (OPLS-DA) followed by combined model scatter plot and analysis of jack-knife confidence levels was performed on the meta probe set level of genes included in the transforming growth factor-β (TGFβ) pathway (26 genes) on tricuspid aortic valve (TAV) and bicuspid aortic valve (BAV) models separately, including dilated and nondilated samples as two discriminant Y-variable groups to identify differentially expressed genes
Given that the TGFβ signaling pathway is believed to be important in aortic aneurysm [6,14], the focus of this study was an exploration of alternative splicing in all genes of this pathway
Summary
Thoracic aortic aneurysm (TAA) is a pathological widening of the aorta resulting from degeneration of the extracellular matrix and loss of smooth muscle cells in the tunica media. Mutations in the TGFβ receptors impair the signaling cascade in other Marfan syndrome–related disorders, including Loeys-Dietz syndrome as well as familial TAA and aortic dissection. The present study was based on further investigation of the importance of alternative splicing in the TGFβ signaling pathway with respect to dilated and nondilated thoracic aorta tissues in patients with TAV and BAV. Our approach to alternative splicing analysis is based on a microarray technology (GeneChip Human Exon 1.0 ST Affymetrix arrays) that measures the expression of single exons to identify alternatively spliced events With this kind of array it is possible to investigate both alternative splicing patterns and differential gene expression. The findings in the present study demonstrate that the two different types of aorta tissues, dilated and nondilated, show different alternative splicing patterns in the TGFβ pathway with respect to TAV and BAV. Diverging alternative splicing patterns observed between the two valve types indicate that the underlying molecular mechanisms of dilatation in TAV patients differ from the mechanisms of dilatation in BAV patients
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