Left ventricular global transcriptional profiling in human end-stage dilated cardiomyopathy

Abstract

We employed ABI high-density oligonucleotide microarrays containing 31,700 sixty-mer probes (representing 27,868 annotated human genes) to determine differential gene expression in idiopathic dilated cardiomyopathy (DCM). We identified 626 up-regulated and 636 down-regulated genes in DCM compared to controls. Most significant changes occurred in the tricarboxylic acid cycle, angiogenesis, and apoptotic signaling pathways, among which 32 apoptosis- and 13 MAPK activity-related genes were altered. Inorganic cation transporter, catalytic activities, energy metabolism and electron transport-related processes were among the most critically influenced pathways. Among the up-regulated genes were HTRA1 (6.9-fold), PDCD8(AIFM1) (5.2) and PRDX2 (4.4) and the down-regulated genes were NR4A2 (4.8), MX1 (4.3), LGALS9 (4), IFNA13 (4), UNC5D (3.6) and HDAC2 (3) (p<0.05), all of which have no clearly defined cardiac-related function yet. Gene ontology and enrichment analysis also revealed significant alterations in mitochondrial oxidative phosphorylation, metabolism and Alzheimer's disease pathways. Concordance was also confirmed for a significant number of genes and pathways in an independent validation microarray dataset. Furthermore, verification by real-time RT-PCR showed a high degree of consistency with the microarray results. Our data demonstrate an association of DCM with alterations in various cellular events and multiple yet undeciphered genes that may contribute to heart muscle disease pathways.

Fig. 1

Heatmap of genes that were significantly modulated due to DCM. Hierarchical clustering clearly separated individuals as either DCM patients or normal controls. Highly expressed genes are indicated in red, intermediate in black, and weakly expressed in green. Only 100 of the most significantly altered genes are shown for readability.

Fig. 2

Functional (A) and canonical pathway (B) analysis of DCM specific genes. X-axis indicates the significance (-log P value) of the functional/pathway association that is dependent on the number of genes in a class as well as biologic relevance. The threshold line represents a P value of 0.05.

Fig. 3

Functional network analysis of DCM specific genes. Top two scoring gene interaction networks with high relevancy scores (significance: score=48) for the DCM specific genes. A score of three indicates that there is 1/1000 (score=−log (P value)) chance that the focus genes are assigned to a network randomly. Green indicates down-regulated, red, up-regulated. The color intensity is correlated with fold change. Straight lines are for direct gene to gene interactions, dashed lines are for indirect ones.

Fig. 4

Confirmation of the microarray gene expression for six randomly selected differentially expressed genes by qRT-PCR. Ratio of expression for each gene in DCM group to normal control (fold change) was log₂ transformed for microarray data and real-time RT-PCR. Dark bars represent microarray hybridizations, and, and grey bars represent values from qRT-PCR. The error bar represents standard deviation (SD) over three experiments.