Genomic and proteomic profiling II: comparative assessment of gene expression profiles in leiomyomas, keloids, and surgically-induced scars

Abstract

Background: Leiomyoma have often been compared to keloids because of their fibrotic characteristic and higher rate of occurrence among African Americans as compared to other ethnic groups. To evaluate such a correlation at molecular level this study comparatively analyzed leiomyomas with keloids, surgical scars and peritoneal adhesions to identify genes that are either commonly and/or individually distinguish these fibrotic disorders despite differences in the nature of their development and growth.

Methods: Microarray gene expression profiling and realtime PCR.

Results: The analysis identified 3 to 12% of the genes on the arrays as differentially expressed among these tissues based on P ranking at greater than or equal to 0.005 followed by 2-fold cutoff change selection. Of these genes about 400 genes were identified as differentially expressed in leiomyomas as compared to keloids/incisional scars, and 85 genes as compared to peritoneal adhesions (greater than or equal to 0.01). Functional analysis indicated that the majority of these genes serve as regulators of cell growth (cell cycle/apoptosis), tissue turnover, transcription factors and signal transduction. Of these genes the expression of E2F1, RUNX3, EGR3, TBPIP, ECM-2, ESM1, THBS1, GAS1, ADAM17, CST6, FBLN5, and COL18A was confirmed in these tissues using quantitative realtime PCR based on low-density arrays.

Conclusion: the results indicated that the molecular feature of leiomyomas is comparable but may be under different tissue-specific regulatory control to those of keloids and differ at the levels rather than tissue-specific expression of selected number of genes functionally regulating cell growth and apoptosis, inflammation, angiogenesis and tissue turnover.

Figure 1

Cluster analysis of 1124 differentially expressed transcripts in leiomyomas (N = 6) form African Americans (AAL1, AAL2 and AAL3), Caucasians (CL1, CL2, and CL3) and in keloids (S3 and S4) and incisional scars (S1 and S2) identified following supervised and unsupervised analysis and p ranking of P < 0.005 followed by 2-fold cutoff change selection (Affymetrix U133A). Genes represented by rows were clustered according to their similarities in expression patterns for each tissue identified as A and B. The dendrogram displaying similarity of gene expression among the cohorts is shown on top of the image, and relatedness of the arrays is denoted by distance to the node linking the arrays. The incisional scar (S1) and keloids were from African American patients. The shade of red and green indicates up- or down-regulation of a given gene according to the color scheme shown below.

Figure 2

Cluster analysis of 206 differentially expressed genes in leiomyomas from Caucasians (CL1, CL2, and CL3) and peritoneal adhesions (A1, A2, A3) using Affymetrix U95 array. The genes were selected based on supervised and unsupervised assessment and p ranking at P < 0.01 followed by 2-fold cutoff change selection. The genes represented by rows were clustered according to their similarities in expression patterns for each tissue and identified as A and B.

Figure 3

The bar graphs show the relative mean expression levels of 12 genes (E2F1, RUNX3, EGR3, TBPIP, ECM-2 ESM1, THBS1, GAS1, ADAM17, CST6, FBLN5, and COL18A1) in leiomyomas (LYM), keloids/incisional scars (Scar) and peritoneal adhesions (P. Adhesion) using realtime PCR and LDA as described in materials and methods section. Values on the y-axis represent an arbitrary unit derived from the mean expression level of these genes in each tissue with their mean expression values in leiomyomas set at 1 independently for each gene prior to normalization against their expression levels in myometrium form a Caucasian serving as control. The asterisks * indicate statistical difference between the expression of these genes with arrows pointing the difference between each group. A probability level of P < 0.05 was considered significant.